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Has anyone recorded directly detected 15N spectra on UnityInova? We have a triple resonance HCN probe. The sample I've used was natural abundance 1M NaNO3 - sodium nitrate. I've tried to set up cable analogously to 13C detection - through the second preamp, but could not see any signal at all. |
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N15 in very difficult to observe using direct detection; the frequency is low, natural abundance is low, its T1 is insanely long, sensitivity is low: 2% of C13. What you need is a sample that can be observed ideally in a single scan. Your 1M NaNO3 is not going to work. When I'm setting up, I use 90% formamide in DMSO-d6. 15N labelled NaNO3 is also available and would be a good choice too. Regards, Tim Thanks, Tim. How do you set the cables? - Evgeny Fadeev (Aug 07 '09 at 14:04) |
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I have used the same cable configuration as for C13 with a broadband probe. 90% formamide in DMSO can be detected in a single scan and gives good S/N in a few scans. On the HCN probe I think the H coil is the innermost coil and so 15N detection is even more difficult than on broadband probes. Also remember 15N has a negative gamma so having the proton decoupler on decreases the 15N signal. Use a sequence that gates the decoupler on during acquisition. On varian S2pul with dm='nny' or on bruker zgig. Tesla |
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On an unlabeled sample, this could be ugly. Even on a 5-10 mM labeled peptide the last person I know who did this with an HCN probe had to signal average for the better part of 2 days to get reasonably good S/N. I've really only done direct 15N observe on concentrated, labeled samples. But, if you're determined to continue: Make sure you use the same transmitter cable from the channel 2 amplifier that you normally use for 13C - all low band nuclei get observed on channel 2 through the low band preamp (tn='N15'). This will probably give you different pulse widths if you normally do 15N on channel 3. The rest of this should be pretty obvious, but I'm including it for completeness: Make sure the "probe" port on the preamp is connected to the 15N connector on the probe. If you move the cable that's normally use for 13C, don't forget to change the barrel filter as well. Make sure you're using the correct 1/4 wave cable on the preamp. If you can, make sure your 15N carrier is set in a reasonable place so you don't have to collect a really wide spectral window. Finally, don't forget to put everything back the way it was when you're finished. |
