title: Nuclear magnetic resonance at the picomole level of a DNA adduct.
abstract: We investigate the limit of detection for obtaining NMR data of a DNA adduct using modern microscale NMR instrumentation, once the adduct has been isolated at the picomole level. Eighty nanograms (130 pmol) of a DNA adduct standard, N-(2'-deoxyguanosin-8-yl)-2-acetylaminofluorene 5'-monophosphate (AAF-dGMP), in 1.5 μL of D₂O with 10% methanol-d₄, in a vial, was completely picked up as a droplet suspended in a fluorocarbon liquid and loaded efficiently into a microcoil probe. This work demonstrates a practical manual method of droplet microfluidic sample loading, previously demonstrated using automated equipment, which provides a severalfold advantage over conventional flow injection. Eliminating dilution during injection and confining the sample to the observed volume produce the full theoretical mass sensitivity of a microcoil, comparable to that of a microcryo probe. With 80 ng, an NMR spectrum acquired over 40 h showed all of the resonances seen in a standard spectrum of AAF-dGMP, with a signal-to-noise ratio of at least 10, despite broadening due to previously noted effects of conformational exchange. Even with this broadening to 5 Hz, a two-dimensional total correlation spectroscopy spectrum was acquired on 1.6 μg in 18 h. This work helps to define the utility of NMR in combination with other analytical methods for the structural characterization of a small amount of a DNA adduct.
authors.affiliation: Barnett Institute of Chemical and Biological Analysis, ‡Department of Pharmaceutical Sciences in the Bouve College of Health Sciences, and §Department of Chemistry and Chemical Biology, Northeastern University , Boston, Massachusetts 02115, United States.
authors.forename: Roger
authors.initials: R
authors.lastname: Kautz
authors.forename: Poguang
authors.initials: P
authors.lastname: Wang
authors.forename: Roger W
authors.initials: RW
authors.lastname: Giese
mesh_terms: D001331:Automation; D018736:DNA Adducts / Q000032:analysis* / Q000592:standards; D005466:Fluorocarbons / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000295:instrumentation / Q000592:standards; D046210:Microfluidic Analytical Techniques; D012015:Reference Standards
chemical_list: D018736:DNA Adducts; D005466:Fluorocarbons
references.citation: Lab Chip. 2008 Feb;8(2):198-220
references.pmid: 18231657
references.citation: Biochem Soc Trans. 2012 Aug;40(4):615-23
references.pmid: 22817704
references.citation: Anal Chem. 2012 Apr 17;84(8):3811-9
references.pmid: 22409256
references.citation: J Biomol Struct Dyn. 1986 Apr;3(5):935-48
references.pmid: 3271419
references.citation: Anal Chem. 1998 Dec 15;70(24):5326-31
references.pmid: 9868921
references.citation: Anal Methods. 2012 Jan 1;4(5):1315-1325
references.pmid: 22707983
references.citation: Curr Opin Chem Biol. 2002 Oct;6(5):711-6
references.pmid: 12413558
references.citation: Biochemistry. 1967 Jan;6(1):177-82
references.pmid: 6030315
references.citation: J Nat Prod. 2010 Sep 24;73(9):1465-9
references.pmid: 20812673
references.citation: J Magn Reson. 2006 Apr;179(2):290-3
references.pmid: 16423543
references.citation: Anal Bioanal Chem. 2012 Jan;402(1):61-8
references.pmid: 21879299
references.citation: Anal Chem. 2008 Nov 1;80(21):8045-54
references.pmid: 18834150
references.citation: Anal Chem. 2004 May 15;76(10):2966-74
references.pmid: 15144211
references.citation: J Comb Chem. 2005 Jan-Feb;7(1):14-20
references.pmid: 15638474
references.citation: Nat Protoc. 2007;2(11):2772-81
references.pmid: 18007613
references.citation: Carbohydr Res. 2011 Oct 18;346(14):2244-54
references.pmid: 21824609
references.citation: J Magn Reson. 2001 Dec;153(2):215-22
references.pmid: 11740897
references.citation: Chem Res Toxicol. 2012 Dec 17;25(12):2737-43
references.pmid: 23113579
references.citation: Curr Opin Drug Discov Devel. 2009 Mar;12(2):197-206
references.pmid: 19333865