JPH10142175A - Measuring method for nuclear magnetic resonance - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a three-dimensional HMBC(heteronuclear multiple band correlation) method in which a spin development time constant is used as a time variable parameter. SOLUTION: Pulses at 90 to 180 deg. are applied to a<1> H nucleus, a prescribed time ▵1 is delayed since the application of the pulses at 90 deg. to the<1> H nucleus, and pulses at 90 to 90 to 90 deg. are applied to a<13> C nucleus. Then, the<1> H nucleus is observed for a detecting time t3 , a second development time t2 is set between second pulses at 90 deg. and third pulses at 90 deg. with reference to the<13> C nucleus during a period in which a first development time t1 is between the first pulses at 90 deg. and the second pulses at 90 deg. with reference to the<13> C nucleus, and a pulse sequence in which the pulses at 180 deg. are applied to the<1> H nucleus is used at a point of time of half the development time t2 .

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to nuclear magnetic resonance (NM)
R) Regarding the measurement method, in particular, HMBC (Heteronuclear Multiple-Bond Corre
lation) Law.

[0002]

2. Description of the Related Art One effective NMR measurement method for elucidating the structure of a natural organic compound using NMR is as follows.
HMBC (Heteronuc
lear Multiple-Bond Correlation) method. FIG. 1 shows H
4 shows a basic pulse sequence of the MBC method. In this pulse sequence, a 90 ° -180 ° pulse is applied to the 1H nucleus, while 90 pulses to the 1H nucleus are applied to the 13C nucleus.
° 90 ° -90 ° with a delay of Δ1 from pulse application
A −90 ° pulse is applied, and observation of the FID signal (free induction decay signal) is performed for the 1H nucleus over the detection time t2. The period between the first 90 ° pulse and the second 90 ° pulse for the 13C nucleus is set to the development time Δ2.

The expansion time t1 is the second 90 ° for the 13C nucleus.
Set between the pulse and the third 90 ° pulse, the deployment time t
At half the time, a 180 ° pulse is applied to the 1H nucleus.
By this 180 ° pulse, the magnetization is developed in the state of two quantum transitions (Double-Quantum Coherence = DQC) in the first half of the development time t1, and in the second half is zero quantum transition (Zero-Quantum Cohence).
erence = ZQC).

The measurement is repeated a predetermined number of times while sequentially changing the development time t1 in small steps, and a series of FID signals (free induction decay signal) obtained as a result is converted to a time t2.
After performing a Fourier transform on the time t1, a HMBC spectrum can be obtained by performing a Fourier transform on the time t1.

In the structural analysis of natural organic compounds, it is important to obtain a good HMBC spectrum.

[0006]

One of the problems of the HMBC method is that spin expansion times Δ1 and Δ2 corresponding to expected spin coupling must be set. If the set values of the spin expansion times Δ1 and Δ2 are inappropriate, no signal is observed. In addition, when a proton to be observed is spin-coupled with many protons to form a broad signal, or when the spin coupling with carbon is small, it is often difficult to observe a cross peak.

Further, broad protons having a short transverse relaxation time T2 are often not observed due to rapid signal decay.

[0008] In view of the above-mentioned points, the present invention relates to a HMBC.
It is an object of the present invention to provide a three-dimensional HMBC method which solves the above problems by expanding the method into three dimensions.

[0009]

In order to achieve this object, the nuclear magnetic resonance measurement method of the present invention uses a 90
° -180 ° pulse is applied, and the development time t1 is delayed from the application of the 90 ° pulse to the 1H nucleus to 90 ° to the 13C nucleus.
A −90 ° -90 ° pulse is applied, observation is performed for the 1H nucleus for a detection time t3, and an expansion time t2 is set between the second 90 ° pulse and the third 90 ° pulse for the 13C nucleus. To the 1H nucleus at half the time t2
° Using a pulse sequence in which a pulse is applied, the measurement is repeatedly performed for a predetermined number of combinations in which the development times t1 and t2 are changed in minute steps, respectively.
The series of FID signals obtained as a result is converted to time t3, t2, t1.
Is characterized in that a three-dimensional NMR spectrum is obtained by performing a Fourier transform on.

[0010]

Embodiments of the present invention will be described below in detail with reference to the drawings. FIG. 2 shows a basic pulse sequence of the three-dimensional HMBC method used in the nuclear magnetic resonance measurement method of the present invention. As can be seen by comparing with the conventional pulse sequence of FIG. 1, Δ2, t1,
t2 has been replaced by new t1, t2, and t3, respectively. The detection of the FID signal is performed during the detection period t3.

The measurement by the sequence of FIG.
The measurement is performed in exactly the same way as the two-dimensional NMR measurement. That is, t1
, T2 are set to appropriate initial values t10, t20, step increments .DELTA.t1, .DELTA.t2, and measurement point numbers N1, N2, respectively. In the measurement, first, t1 is fixed at t1 = t10, and t2 is calculated from t2 = t20 to (N2 -1) .DELTA.t2.
The same measurement is performed over N2 steps, changing at steps of .DELTA.t2 up to .DELTA.t2.
The process is performed for each of the N1 steps while changing in increments of .DELTA.t1 until t1, so that a total of N1.times.N2 measurements constitutes one three-dimensional NMR measurement. If necessary, this is repeated a plurality of times and integrated, thereby improving the SN ratio.

The N1.times.N thus obtained is
Set data D (t1, t2, t3) of two FID signals
Is converted from the time domain to the frequency domain by performing a Fourier transform on t3, t2 and t1 using a computer, and the three-dimensional NMR spectrum data S (F1, F2
2, F3) are obtained.

In the three-dimensional NMR method, a complicated cross peak appearing in a two-dimensional NMR spectrum is developed in a three-dimensional space, so that the spectrum can be easily analyzed. In the three-dimensional HMBC spectrum obtained by the measurement based on the pulse sequence in FIG. 2, as shown in FIG.
A three-dimensional NMR spectrum is obtained in which the axis is proton, the F2 axis is carbon, and the F3 axis is proton. Further, as shown in FIG. 3B, in the slice spectrum of the proton HA on the F1 axis, an HMBC spectrum from the proton HA is obtained.
Similarly, in the slice spectrum of the proton proton HB, an HMBC spectrum from the proton HB is obtained.
It can be seen that the complex two-dimensional NMR spectrum has been greatly simplified. FIG. 3 (c) shows the F2 / F3 axis projection spectrum.

Example 1 FIG. 4 shows the structural formula of a port sewing machine used as a sample, and FIG. 5 shows a three-dimensional HMBC projection spectrum obtained by the NMR measurement method of the present invention. FIG. 5 (a) shows the F1 / F3 axis projection spectrum, and FIG. 5 (b) shows the F2 / F3 axis projection spectrum. The measurement conditions are
F1 (1H) × F2 (13C) × F3 (1H) = 2200 × 14
00 × 2200Hz, measurement point is 64 × 150 × 2
The measurement was performed for about 8 hours using the pulse field gradient method with 56 times of integration.

In the F1 / F3-axis projection spectrum shown in FIG. 5 (a), only the chemical shift between protons and protons is observed on the diagonal axis. On the other hand, F2 / F3 axis projection spectrum and F1 / F3
In the F2 axis projection spectrum, ordinary HMB
A spectrum similar to the C spectrum is obtained.

FIG. 6 shows a three-dimensional HMBC of a port sewing machine.
The F2 / F3-axis projection expanded spectrum obtained from the spectrum (FIG. 6 (a)) and the slices of protons of the F1-axis from 24Me, 6Me and 18Me (FIG. 6).
(B) to (d)) are shown. It can be seen that the slice spectrum is a simple spectrum with good separation compared to the projection spectrum. Therefore, analysis can be easily performed.

Normally, when the observation region of the F1 axis is selected by three-dimensional NMR, if it is a proton, a step increment at which the chemical shift of the proton can be grasped (Δt1 = 200 μsec.about.50)
(0 μs) and acquire a three-dimensional NMR spectrum. In this case, the effect of the J modulation cannot be shifted to the F2 axis without acquiring a large number of data points.

On the other hand, the present inventors ignored the chemical shift in the observation region of F1 and attempted to develop as few as 16 to 32 data points at Δt1 = 5 to 10 msec. As a result, the obtained spectrum becomes a small matrix spectrum with few data points, and more effectively,
It was confirmed that the effect of the ng-range J modulation can be shifted to the F2 axis.

Example 2 FIG. 7 shows the structural formula of monazomycin used as a sample, and FIG. 8 shows Δt based on the above-mentioned concept.
The three-dimensional HMBC projection spectrum obtained with the setting of 1 is shown. FIG. 9 also shows a conventional HMBC spectrum for comparison. The spectrum of FIG. 8 is obtained by F1 × F2 × F3 = 125 × 23000 × 2900.
Hz, number of measurement points 16 × 256 × 512, t10 = 20
The measurement was performed for m seconds, Δt1 = 4 msec, and the required measurement time was about 7 hours. The spectrum in FIG. 9 was measured at the same time with Δ set to 60 ms.

In the HMBC spectrum shown in FIG.
No cross peak was observed from the broad signals at positions 46 and 11. On the other hand, in the spectrum of FIG. 8, the proton at the 18th position is 18Me, 19, 1
Cross peaks of 7 and 16 are observed, and cross peaks of 46 Me, 44, 45 and 47 are observed from the proton at position 46. Although no cross-peak is observed for the 11-position proton in the spectrum of FIG. 9, 9, 10, 12, and 13 are observed from the 11-position proton in the spectrum of FIG. Become. In particular, it is often difficult to prove the bond relationship between methylene and methylene with a complex compound because the transverse relaxation time T2 of the signal is short and the spin is complicatedly split. You can find relationships.

FIG. 10 shows the proton at position 18 in FIG.
10 and slice data cut out from the spectrum of FIG. 9. In the slice data from the normal HMBC spectrum in FIG. 10B, only a weak cross peak is observed, but in the slice data from the spectrum in FIG. 8A, the signal intensity is extremely increased. You can see that there is. In the ordinary HMBC method, the spin expansion time Δ
Is set to one value (usually about 60 msec), and no signal that does not conform to this value is observed.

Although it is difficult to set the value of Δ to the optimal value of various spins by the ordinary HMBC method, the three-dimensional HMBC method of the present invention eliminates the problem of setting the spin expansion time. Can be.

[0023]

As described in detail above, according to the present invention,
Since the three-dimensional HMBC method using the spin expansion time constant in the HMBC method as a time variable parameter has been realized, it is possible to eliminate the problem of setting the spin expansion time, and by extracting a projection spectrum from the obtained three-dimensional HMBC spectrum. 2. A spectrum that enables analysis as a two-dimensional NMR spectrum is obtained.

[Brief description of the drawings]

FIG. 1 is a diagram showing a basic pulse sequence of the HMBC method.

FIG. 2 shows 3 used in the nuclear magnetic resonance measurement method of the present invention.
FIG. 3 is a diagram showing a basic pulse sequence of the dimensional HMBC method.

3 is a schematic diagram showing a three-dimensional HMBC spectrum, a slice spectrum, and a projection spectrum obtained by measurement based on the pulse sequence of FIG. 2;

FIG. 4 is a view showing a structural formula of a port sewing machine.

FIG. 5 shows a three-dimensional HM obtained by the NMR measurement method of the present invention.
It is a figure which shows a BC projection spectrum.

FIG. 6 is an enlarged F2 / F3-axis projection spectrum obtained from a three-dimensional HMBC spectrum of a port sewing machine, and 24Me, 6Me, and 18Me of protons on the F1 axis.
FIG. 6 is a diagram showing slices from a.

FIG. 7 shows the structural formula of monazomycin.

FIG. 8 shows a three-dimensional HMBC projection spectrum of monazomycin.

FIG. 9 is a diagram showing an HMBC spectrum of monazomycin obtained by a conventional HMBC method.

10 is a diagram showing slice data cut out from the spectra of FIGS. 8 and 9 for the proton at position 18. FIG.

Claims (2)

[Claims] 1. A 90 ° -180 ° pulse is applied to a 1H nucleus, and a 90 ° -90 ° -90 ° pulse is applied to a 13C nucleus by delaying a predetermined time Δ1 from the application of a 90 ° pulse to a 1H nucleus. And the observation is performed for the 1H nucleus over the detection time t3, and the first development time t1 is set to the first time for the 13C nucleus.
During the period between the 90 ° pulse and the second 90 ° pulse, the second deployment time t2 is determined by the second 90 ° pulse and the third
A predetermined number of combinations in which the expansion times t1 and t2 are changed in minute steps, respectively, using a pulse sequence that is set during the 90 ° pulse and the 180 ° pulse is applied to the 1H nucleus at half the expansion time t2 A method for measuring nuclear magnetic resonance, characterized in that a three-dimensional NMR spectrum is obtained by performing a Fourier transform on a series of FID signals obtained as a result at times t3, t2 and t1. 2. The step change increment Δt1 of the development time t1.
The nuclear magnetic resonance measurement method according to claim 1, wherein is set to 5 to 10 msec.