It is often difficult to unambiguously determine structures of molecules, which have many non-protonated carbon atoms. Because such compounds contain quaternary carbons, lack protons and bonding to quaternary carbons each other, HMBC (Heteronuclear Multiple Bond Correlation) experiment cannot provide long-range 1H-13C connectivity. Therefore, INADEQUATE (Incredible Natural Abundance DoublE QUAntum Transfer Experiment), which is 13C-13C correlation experiment at natural 13C abundance, represents a powerful tool for proton-diluted compounds. On the other hand, INADEQUATE is about 200 times less sensitive measurement compared to ordinary 13C measurement in theory. For this reason, INADEQUATE experiment usually requires highly concentrated samples and long experimental times. Experimental time and/or sample amount can be greatly reduced by the use of SuperCOOL probe as shown below.
How to analyze INADEQUATE spectra
13C-13C correlations in INADEQUATE spectra are observed as pairs of doublets symmetrical with respect to the diagonal line of slope 2 Fig.1 shows a correlation between carbons CB and CE. Each signal is split into a doublet by the coupling constant 1J(CBCE).
Fig.1: Correlation pattern typical of INADEQUATE spectra
Application of INADEQUATE
Genistein is a soy-derived isoflavone and has antioxidant activity. The structure contains eight quaternary carbons and some of these quaternary carbons are bound to others. The INADEQUATE spectrum is shown in Fig. 2. The spectrum was collected on a sample containing 16 mg of Genistein using a SuperCOOL probe at 600 MHz. All one-bond 13C-13C correlations were detected within 18 hours.
Fig.2: INADEQUATE spectrum
console: JNM-ECZ600R, SuperCOOL probe
sample: 100mM(16mg) Genistein/DMSO-d6
measurement time: 18h(standard tube)
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