Chemistry Reference
In-Depth Information
precessions at frequencies
J /2 relative to the rotating frame. Because this frame is
significantly tilted from the rotating frame, application of frequent 1 Hpulsesresultsin
incomplete cancelation of the cross term. Therefore, infrequent 1 H pulses are applied
to suppress DD/CSA correlation when the off-resonance effect is significant [ 38 , 39 ].
Other parameters that have to be specifically considered in the R 1 r experiment
are the spatial homogeneity of the B 1 field strength and linearity of the power
amplifier. Although B 1 homogeneity has been improved in recent NMR probes,
estimation of the inhomogeneity remains important to confirm the accuracy of the
obtained R 1 r . For this purpose, measurement of the B 1 inhomogeneity using inverse
detection is useful [ 40 ]. Amplifier linearity has also improved. However, since
pulse power is switched for spin-lock, it is important to check that a phase shift
accompanies a power change, and make a correction should one be needed.
2.1.2 Carr-Purcell-Meiboom-Gill R 2 Experiment
In CPMG R 2 , limitation of applicable B 1 field strength is in general smaller than that
of the R 1 r experiment. Since CPMG 180 pulses are applied with interpulse delays
(2
than that of spin-lock can be
applied. However, in a protein, the RF field strength used in the 15 NCPMG R 2
experiment must be carefully considered because the interpulse delay is set short
(typically, 2
t CP ), CPMG pulses with much stronger
g N B 1 /2
p
t CP ~1 ms) to suppress generation of antiphase terms, N X , Y H Z ,causedby
1 H- 15 N J coupling. For example, if 6 kHz CPMGpulses are appliedwith 2
1ms,
the RF power delivered to the probe is nine times stronger than that of a 2 kHz spin
lock. Given that the duty cycle is ca. 10% in the CPMG experiment, about the same
amount of energy is deposited in the probe in both CPMG and R 1 r experiments.
In CPMG R 2 , off-resonance error is negligible at low magnetic field strength but
significant at high magnetic field strength [ 41 , 42 ]. Although the stronger
t CP ¼
for
each CPMG pulse inverts magnetization more uniformly than the spin-lock, CPMG
pulse train accumulates error caused by a combination of pulse imperfections and off-
resonance effects. The CPMG error of a signal located at off-resonance frequency
f off ( f off is the difference between the signal and carrier frequencies) is estimated
with a function of 2
g N B 1 /2
p
t CP and B 1 . Importantly, the off-resonance error is maximized
at 2
t CP f off ¼
n ( n is an integer): when 2
t CP ¼
1 ms, the off-resonance error is signi-
ficant at f off ¼
1, 2, and 3 kHz [ 41 ]. Magnitude of the error depends on the B 1 field
strength the 180 pulses. This relationship indicates that R 2 canberecordedwithout
significant off-resonance error at 61 MHz for signals because the entire chemical
shift range for amide backbone 15 N signals spans approximately less than
15 ppm.
t CP is used at 15 N 91MHz resonance frequency, signals located
However, if the same 2
at
11 ppm off-resonance suffer from significant errors in measured R 2 values.
It is a disadvantage of CPMG R 2 experiment that there is no simple equation to
correct for CPMG R 2 off-resonance effects. In practice, it is recommended to
discard R 2 data obtained at f off ¼
t CP frequency, and record the data at two
different carrier frequencies. As an alternative, a phase cycle to average out the off-
resonance effect in CPMG R 2 may be used [ 43 , 44 ]. Using this method, R 2 is
n /2
Search WWH ::




Custom Search