NMR

AVANCE NEO-400 WB (Bruker)

bpscherf — Thu, 23 Feb 2023 11:56:43 +0000

AVANCE NEO-400 WB (Bruker) bpscherf Thu, 02/23/2023 - 11:56

A 400WB NMR spectrometer equipped with MAS unit for solid state NMR studies.

This 3-channel UltraShield NMR spectrometer is equipped with the following MAS probes:

4 mm DR-(low range) DVT probe for solid state NMR with wide bore (WB) magnets.
Features of this top-loading double resonance X/H 4mm CP-MAS DVT probe:
- X tuning range from 13C to 109Ag
- High power 1H decoupling
- Max. rotation frequency: 15 kHz
- Active sample volume: 50 ul
- Temperature range: -130°C to 150°C
1.3 mm DVT multinuclear double resonance 15N-31P/19F-1H probe (1.7 microL active volume, 67 kHz spinning speed, VT range -30°C to 70°C)
2.5 mm top-loading DVT multinuclear triple resonance X/Y/19F-1H (8 microL (14 fir thin-walled rotors) active volume, 35 kHz spinning speed, VT range -50°C to 80°C)
4 mm top-loading DVT multinuclear triple resonance X/Y/19F-1H probe (50 microL active volume, 15 kHz spinning speed, VT range -130°C to 150°C)

Unit

NMR

Contacts

Dr. Tali Scherf

Wolf building

Short Spectrometer Manuals

nyhofit — Tue, 08 Nov 2022 10:26:08 +0000

Short Spectrometer Manuals nyhofit Tue, 11/08/2022 - 10:26

These notes include a general introduction to Bruker TopSpin software, and a step-by-step guide to the acquisition and processing of 1D spectra. It is by no mean a replacement for the short training course each user is obliged to take.

The Linux/UNIX operating system

The new spectrometers make use of the Linux operating system. It is a multi-user system, which means that each user has his own ‘account’ and is required to logon to the instrument using its own user name ( userid ). Each userid is password protected, so that only you should be able to logon to your account. The concept of user accounts is a very important one in Unix systems as this defines who created and "owns" the files, as well as where they are to be found and who has permission to alter and/or delete them. The NMR data sets you create (measure) are stored in directories and sub-directories, and each spectrometer user has his own directory in which his FID’s and spectra are to be found. For example, any data created by userid
‘tali ’ will be stored on the disk under the directory: /DISK/data/tali/nmr

(where /DISK specifies the particular disk partition on which the data is kept, e.g. /opt/topspin/data/tali/nmr).

Another important point is that Linux commands are case sensitive , and this therefore applies to commands you may use in TopSpin. Thus ft and FT will not be the same. Lowercase commands should be used at all times in TopSpin.
In the description on the following pages, text you should type is shown in bold italic, and responses from the system are shown in bold .
Getting started & Exiting: You should start at a monitor with the login screen. If this is not the case, exit (logout) as described later. Double click on your userid icon, and enter your password. Open a Unix shell and type topspin to start the program. To exit the program when you finished your session, type exit and confirm. This will return you to a Unix shell. To logout from Unix, use the right mouse button.

Interacting with the TopSpin program

Interaction with the TopSpin program is either by clicking on the icons with the mouse, by selecting a command from the pull-down menus or via keyboard commands (typing the same commands available in the pull-down menus). [A list of common Bruker commands and some basic nomenclature is available under the Instruction Manuals & FAQ section]. In addition, the mouse (left, right and middle) is used to interact with the spectrum directly. The basic TopSpin display (for 1d spectra) which you will usually get when you run the program is shown in the figure below. Other windows, which display other features (lock display, temperature control etc) may also appear at various stages. The main areas of the Main Display shown below include:

Pull-down menus, at the top, that contains the File, Acquire, Process, Analysis, Display, Window and the Help pull-down menus.
Identifier, which is the text that shows the data set currently in use, in the format of data set name (NAME e.g. btx1_g7_303_8), experiment number (EXPNO e.g. 1), processing number (PROCNO e.g. 2), disk partition (DISK e.g. /opt) and user name (USER e.g. tali).
Spectral window, the area used to display the FID and the spectra.
Icon panels (at the left), controls the display of the FID and the spectra at the spectral window (the size, range etc), as well as some additional subroutines.
Command line, just below the spectral window, where you could type the commands (starts with a >).
Information panel (below the command line), at which the current process or the development of the process is being described (e.g. fp: finished).

Data Acquisition

Defining data sets

All data generated by the spectrometer are automatically saved to disk and stored in your directory. Therefore it is always necessary to define a name of your data before doing anything else; this should always be the first thing you do when you start a session on the spectrometer. There is a real chance of you overwriting a previous data set if you fail to do this. To define your new data set, type the command edc and complete the six variable entries (data set name, experiment number, processing number, disk partition, user name and the data type that is obviously NMR).

Reading and editing parameters

Parameters are stored in four separate categories for acquisition, processing, plotting and output devices. All four may be loaded simultaneously with the rpar command. This will display a table of available parameter sets through which you may scroll and select your desired parameters. [Note that different probes may have e.g. different protons parameters so there is no universal parameter set for proton acquisition]. The second table asks which of the four parameter groups you wish to load; it’s better to Copy All .
Very important information included in the parameter sets defines the routing of the rf through the spectrometer. Thus, it is not necessary to change the hardware set up, such as cables and pre-amplifier boxes, when you change experiment or nucleus. Thus, it is possible to change the rf routing via the command edasp. [Try to avoid using it as a new user].
The parameters can be edited with the commands eda (edit acquisition), edp (edit processing), edg (edit graphics i.e. plotting), and edo (edit output device i.e. plotter). At this point it is necessary to point out two different parameter classes: “Set parameters”, are those you enter for use during acquisition, processing etc and can be viewed with eda, edp, edg etc. “Status parameters” that actually relate to the data already acquired or processed and can be seen only with dpa dpp , or dpg . For example, suppose you were to set initial parameters in eda to acquire 64 scans, but you halted acquisition after only 32 scans. The eda table will still indicate 64 scans, as would the parameter ns (number of scans) although this is clearly incorrect for the data you have acquired. However, the dpa table would show ns = 32, the actual number of scans for the data set.

Setting the temperature

within the probe is done using the edte command. It is highly recommended to set the temperature before you insert your sample.

Important acquisition parameters

In order to interact with data acquisition you should go from the main display (used for interacting with processed data) to a similar display for acquisition by selecting from the pull-down menu Acquire -> observe FID window (in most of the spectrometer, the alias a given from the command line, exist). The return icon will always take you back to the main display. The pulse program you will be using contains a list of commands regarding rf pulses, delays etc. The name of the pulse program, as well as of the different acquisition parameters can be edited using the eda command. A reduced list of parameters to be used by the selected pulse program is obtained via the ased command.
- PULPROG- the pulse program you wish to run (e.g. zg, zgpr, noesygrtp19 etc).
- D1- the relaxation delay between scans (sec).
- P1- the 90 deg pulse length, hard pulse (µsec).
- PL1- the power level, the power attenuation of the transmitter pulse (in dB).
- SW- the spectral width (ppm). Typically, in the range of 10-12 ppm for 1.
- SFO1- the transmitter frequency in MHz. Could be set by setting the frequency at the center of the spectral window (O1, in Hz). For aqueous samples, it is usually set on the water resonance.
- TD- the time domain data size. Typically 16-32k points.
- NS- the number of scans to be acquired.
- DS- the number of dummy scans before data is collected, to obtain steady-state (2-16).
- RG- the receiver gain, could be set automatically by the rga command.
Locking and Shimming

To see the lock display go in the pull-down menu: Windows -> lock display. If you have changed the probe, the solvent or the type of sample tube, you have to recall an appropriate set of shim parameters by typing rsh. A list of saved shim files will appear from which you should select the appropriate one. A digital lock system is used on most of the instruments. To lock on the deuterium signal of your solvent, type lock and select your solvent from the list. The lock power, gain and phase should be set correctly automatically. The shim file you have recalled using rsh will not update them, but you can still view them using the vish command. Shimming is done via the shim keyboard located near the workstation keyboard. Further information regarding shimming procedures can be found under the Instruction Manuals & FAQ section.

Tuning the probe

Prior to acquiring the data, the probe must be tuned and matched to optimize its performance at the frequency of the studied nucleus (defined via eda or edasp ) and to maximize the power that is transmitted to the coil. To start the procedure type wobb (in most instruments you can use the alias ) and after ~30 seconds enter the acquisition display ( acqu ). Approach the probe, and using the tuning and matching rods or sliders tune and match the probe. The results are shown both on the LED display of the preamp as well as in the acquisition submenu of TopSpin. A well tune probe will show a signal exactly at the center, with its minima (dip) as deep as possible; a minimum number of illuminated LEDs will further support the quality of the wobbling. When tuning is complete, click on the stop icon, type ii (initial interface) and wait for the routine to finish. When tuning and matching a probe with multiple resonant circuits, it is best to tune and match the lowest frequency circuit first. Thus when wobbling a probe for both 1H and 13C, first do the 13C and then adjust the 1H. When tuning and matching a nucleus that its routing does not go through the preamp, it is required to physically change the cable connections so it would pass through the preamp. Please ask the NMR staff is you are not sure how to do it. After tuning & matching the probe, check once more if further improvement in the shimming is possible.

Calibrating the pulses

. In case you want to calibrate your pulses, or use previously-calibrated values, please check the Pulse Calibration Procedures , as well as the Calibrated Pulse-Length Values .

Acquiring the data

To start data acquisition, enter zg. To see the acquired FID enter the acquisition display (acqu). As long as the acquisition is in progress, information regarding the number of scans, the actual number of scans that have already been acquired and the time remaining till the end of the experiment, will appear on the screen. To terminate acquisition immediately use the stop command, but be aware that it will not save your data (which will be lost). To stop acquisition and view the scans acquired so far, use halt , which will write your data at the end of the current scan and allow processing. If you want to view the data acquired till now, without terminating the acquisition process, use the transfer command, tr . This forces the data from the computer memory (where it is held during the acquisition) onto the disk. Go back to the Main Display by pressing the return icon.

Important note

When leaving a long experiment to run, NEVER leave it on the acquisition window- ALWAYS return to the main menu.
Procedures for calibrating pulse lengths can be found under Instruction Manuals& FAQ.

Data Processing

Once you have your data on disk, you may process it and interact with the result using the Main Display. The parameters that define the processing may be edited with the edp command that will open them in a table.

Zero Filling (ZF). In case no zero filling is required, the size of the spectrum (SI) should be set to half the size of the time domain data, the FID (TD). Setting SI=TD means that zero filling will occur.

Window Functions. In order to increase sensitivity or resolution of the spectrum, we often apply a suitable window function to the FID before the Fourier Transformation. Sensitivity enhancement is done by multiplying the FID with an exponential weighting function, which forces the end of the FID towards zero. This exponential multiplication, done via the command em (prior to the ft ), improves the S/N ratio, but leads to some line broadening as well. The sensitivity enhancement is controlled by the parameter lb which defines the resulted line broadening, in Hz. [It is possible to use the ef, which does the two operations ( em + ft ) consecutively]. For resolution enhancement, one can use the sine-square window, qsin , along with the parameter ssb that defines its shape. One can adjust the parameters for window functions interactively, using the menu Process -> Manual window adjust .

Fourier Transformation (FT). To produce a spectrum (frequency domain data) from the time domain data (FID), the raw data must be Fourier transform by the command ft. The spectrum is saved with the processing number defined with edc, and the raw FID also remained (unchanged) on the disk, and could be viewed using the FID icon.

Phase Correction. After the FT the spectrum does not usually appear normally, it is not a pure absorption spectrum. Zero (non-frequency dependent) and first order (frequency dependent) phase-distortion appears. These may be corrected automatically or manually. The automatic correction is done via apk. The manual correction is done by clicking on the phase button. Usually, one selects the largest peak to define zero-order phase correction by clicking biggest . Now position the cursor over the PH0 icon, press the left mouse and drag to correct the phase of the selected peak (dotted vertical line). When done, repeat the procedure for the PH1 icon and correct all other peaks in the spectrum. In order to terminate the process, click on the return icon, save and return to store the new phase. Spectra can be processed identically afterwards using the command pk, or fp that combines ft + pk . [In cases you want to define another peak- not the largest, manually, define it with the cursor icon, or do it manually without definition].

Spectrum Calibration. Defining the origin of the spectrum can be done automatically using the menu Analysis -> sref. Manual calibration is done using the calibration icon, and selecting the top of the reference peak with the middle mouse. Type then the desired reference valuein ppm, in the popping window.

Base-Line Correction. Distortion in the base line is observed when the noise in the signal-free regions of the spectrum are not scattered around the zero line, but around a polynomial or some other curve. Automatic base line correction is done with the abs that subtracts a polynomial from the spectrum. The manual correction is done using the menus Process -> Special Processing -> Baseline correction . Five polynomial functions (A to E) should be adjusted, by clicking and dragging on each. Start with A (linear offset) and go through till E. When suitable fit has been found, click on diff to see the result. To save values use return , then save and return . [You can do FID base line correction prior to the window function and FT, by using the command bc].

Integration. To enter the manual integration routine click on the integrate icon. Place the cursor over the spectrum and click with the left mouse button: the cursor will slide along the spectrum (left mouse terminates this). Go along the spectrum and use the middle mouse to mark the ‘start’ and ‘finish’ of each integral region, after which each new trace is displayed. The icon calibrate can be used to give a numerical value to a specific (current) integral trace, while all others will be scaled accordingly, so the figures will not have an arbitrary value. To define the ‘current/active’ integral trace, click on the left mouse, moves the cursor to under the required trace and click the left button again. An arrow will indicate that this is the ‘current/active’ trace, and all buttons under current will apply to this trace. Now calibrate this trace. Integrals can be saved via return, and save as ‘intrng’ & return . The list of integrals can be printed with li (or may be included in the plot of the spectrum).

Peak Picking. In order to avoid picking noises, it is recommended to set the minimum threshold for the picking procedure. First ensure that the Y-axis units are in centimeters (use the Y icon). Select the utilities button and click on MI. Move the white horizontal line and define the minimum level (MI in cm) using the left mouse. The maximum cut off may also be defined using the MAXI button and repeating the procedure. To view all peaks you have picked on screen, type pps (and plot them using the print button).

Other Useful Tasks

Title. A title may be set using the command setti (use the alias t).
Dual display of two spectra: use the command edc2 to define the second data set. Press the icon dual to display the current and the ‘second’ data set, one on the top of the other.

A brief summary of the steps

Login to the system (your account)
Run topspin
Define your data set using edc (read appropriate parameters if required using rpar)
Set the temperature within the probe (edte)
Insert your sample
Recall a shim file (rsh), lock and shim (BSMS keyboard)
Tune and match the probe (wobb or atmm)
Calibrate the pulses if required
Set acquisition parameters (eda)
Acquire the data (zg)
Process the FID (window, ft, bc, peak picking, integr etc).

Pulse Program Information and Properties

renad — Mon, 07 Nov 2022 14:19:04 +0000

Pulse Program Information and Properties renad Mon, 11/07/2022 - 14:19

The nomenclature of parameters as described in Pulprog.info.

For a pulseprogram the first characters (usually up to 6, but sometimes more) specify the type of experiment, e.g. DEPT, COSY,NOESY etc..
Further properties of the pulseprogram are indicated by a two-character code, which is added to the name in alphabetical order.

For 2D experiments the mode (absolute value,phase sensitive or echo-antischo) is always indicated. H- or X-decoupling is assumed to be default for heteronuclear experiments,but not for homonuclear ones (except inad).
In case of redundant information some two-character codes may be ommitted.

The two-character codes used are the following

ar     experiment for aromatic residues
at     adiabatic TOCSY
bi     with bird pulse for homonuclear J-decoupling
bp     using bipolar gradients
cc     cross correlation experiment
cp     with composite pulse
ct     constant time
cw     decoupling using cw command
cx     using CLEANEX_PM
dc     decoupling using cpd command
df     double quantum filter
di     with DIPSI mixing sequence
dh     homonuclear decoupling in indirect dimension
dw     decoupling using cpd command only during wet sequence
dq     double quantum coherence
ea     phase sensitive using Echo/Antiecho method
ec     with E.COSY transfer
ed     with multiplicity editing
es     excitation sculpting
et     phase sensitive using Echo/Antiecho-TPPI method
fb     using f2 - and f3 - channel
fd     using f1 - and f3 - channel (for presaturation)
fr     with presaturation using a frequency list
ft     using f1 -, f2 - and f3 - channel (for presaturation)
fh     F-19 observe with H-1 decoupling
fp     using a flip-back pulse
fl     for F-19 ecoupler
f2     using f2 - channel (for presaturation)
f3     using f3 - instead of f2 - channel
f4     using f4 - instead of f2 - channel
gd     gated decoupling using cpd command
ge     gradient echo experiment
gp     coherence selection using gradients with ":gp" syntax
gr     coherence selection using gradients
gs     coherence selection using shaped gradients
hb     hydrogen bond experiment
hc     homodecoupling of a region using a cpd-sequence
hd     homodecoupling
hf     H-1 observe with F-19 decoupling
hs     with homospoil pulse
ia     InPhase-AntiPhase (IPAP) experiment
ig     inverse gated
ii     using inverse (invi/HSQC) sequence
im     with incremented mixing time
i4     using inverse (inv4/HMQC) sequence
jc     for determination of J coupling constant
jd     homonuclear J-decoupled
jr     with jump-return pulse
lp     with low-pass J-filter
lq     with Q-switching (low Q)
lr     for long-range couplings
l2     with two-fold low-pass J-filter
mf     multiple quantum filter
ml     with MLEV mixing sequence
mq     using multiple quantum
nd     no decoupling
no     with NOESY mixing sequence
pc     with presaturation and composite pulse
pg     power-gated
ph     phase sensitive using States-TPPI, TPPI, States or QSEC
pl     preparing a frequency list
pn     with presaturation using a 1D NOESY sequence
pp     using purge pulses
pr     with presaturation
ps     with presaturation using a shaped pulse
qf     absolute value mode
qn     for QNP-operation
qs     phase sensitive using qseq-mode
rd     refocussed
rl     with relay transfer
rs     with radiation damping suppression using gradients
ru     using radiation damping compensation unit
rv     with random variation
r2     with 2 step relay transfer
r3     with 3 step relay transfer
se     spin echo experiment
sh     phase sensitive using States et al. method
si     sensitivity improved
sm     simultaneous evolution of X and Y chemical shift
sp     using a shaped pulse
sq     using single quantum
ss     spin-state selective experiment
st     phase sensitive using States-TPPI method
sy     symmetric sequence
s3     S3E experiment
tf     triple quantum filter
tp     phase sensitive using TPPI
tr     using TROSY sequence
tz     zeroquantum (ZQ) TROSY
ul     using a frequency list
us     updating shapes
wg     watergate using a soft-hard-soft sequence
wt     with WET watersuppression
w5     watergate using W5 pulse
xf     x-filter experiments
xy     with XY CPMG sequence
x1     x-filter in F1
x2     x-filter in F2
x3     x-filter in F3
zf     with z-filter
zq     zero quantum coherence
zs     using a gradient/rf spoil pulse
1d     1D version
1s     using 1 spoil gradients
11     using 1-1 pulse
19     using 3-9-19 pulse
2h     using 2H lockswitch unit
2s     using 2 spoil gradients
3d     3D sequence
3s     using 3 spoil gradients
30     using a 30 degree flip angle
45     using a 45 degree flip angle
90     using a 90 degree flip angle
135    using a 135 degree flip angle

Typical experiment names would be:
cosy, dept, dipsi2, hmbc, hmqc, hoesy, hsqc, inad, inept, mlev, noesy, roesy or trosy.

Inverse correlations are denoted as hmbc, hmqc or hsqc.
Experiments with a BIRD sequence in the beginning also contain a bi in the name.

1D experiments, which are analogues of 2D experiments by virtue of a selective pulse, start with sel.

Semiselective 2D experiments have the same name as the unselective version but with an s at the beginning:
scosyph <-> cosyph.

A phase-sensitive (States-TPPI, TPPI etc.) NOESY experiment with presaturation would then be:

noesy + ph + pr = noesyphpr.

In the other direction the pulseprogram hmbcgplpndqf would be

hmbc + gp + lp + nd + qf

and therefor an:

inverse correlation for long-range couplings (HMBC) with coherence selection using gradients with ":gp" syntax,
low-pass J-filter,
no decoupling
in absolute value mode.

Comments like:

   ;avance-version
   ;begin ____
   ;end ____

with (____ = MLEV17, DIPSI2, ...)

are evaluated by NMRSIM for the pulseprogram display and should therefor not be removed. The syntax for begin/end statements allows characters, numbers and '_'. Arithmetic operators must not be used.

The comments:
;preprocessor-flags-start
;preprocessor-flags-end

are also evaluated to identify flags used in the pulseprogram and must also not be removed.

Common Bruker Commands and Nomenclature

renad — Mon, 07 Nov 2022 14:18:06 +0000

Common Bruker Commands and Nomenclature renad Mon, 11/07/2022 - 14:18

edc- edit current data set (can be used to copy current data set parameters into a new name)
eda- edit acquisition parameters
edp- edit processing parameters
edo- edit output device (printer/plotter)
edg- edit graphics, the plotting parameters
acqu [a]- go to the acquisition menu
ased- edit acquisition parameters relevant to the current pp
edasp- edit the rf routing (the channels)
wobb [w]- starts the frequency sweep of the tune and match of the specified (e.g. 1H) circuit of the probehead.
atmm- (to be used for those probes equipped with Automatic Tuning & Matching, ATM)- enables maual adjustment of the wobbling (tuning & matching) of the probehead circuit
atma- (to be used for those probes equipped with Automatic Tuning & Matching, ATM)- enables automatic adjustment of the wobbling (tuning & matching) of the probehead circuit
d- indicates delays (d1 to d31), given in seconds, and can be given as a parameter in eda/ased or can be written explicitly in the pulse program (e.g. 20µ, 15m, “d4=d2+p13*2-2µ”).
p- indicates rectangular pulses (p1 to p31) which are given in µsec.
sp- indicates shaped pulses (sp1 to sp31) given in µsec, and require additional definitions of spnam (the name of the sp), and spoff (the offset of the pulse).
pl- indicates powerlevels (pl1 to pl31) given in dB units (attenuation), and are set on a specific channel (e.g. pl2:f2).
o1- the carrier frequency offset (usually) of the 1H pulses, in Hz
o1p- O1 in ppm units
sfo1- the spectrometer frequency (SF) + O1, in MHz
sw- total spectral width, in ppm units
swh- the equivalent of SW, given in Hz units
td- time domain size of the acquired FID
si- size of the frequency-domain spectrum
ns- number of scans (depends on the phase cycling of the pulse program)
ds- number of dummy scans
rg - the receiver gain, can be set automatically by rga
zg- start running the currently-defined pulse program (regardless of its name).
expt- experiment time
ft- the Fourier transformation to the acquired FID
pk- phase correction using previously set parameters (PH0, PH1)
fp- performs consecutively ft + pk
qsin- a sine square window function applied to the FID before ft, controlled by the parameter ssb
ssb- square sine bell parameter used by qsin
em- exponential multiplication of the FID, controlled by lb
lb- the line-broadening parameter used by em, given in Hz
ef – performs consecutively em + ft
efp- performs consecutively em + ft + pk
setti [t]- sets the title one can give to the data set
gradients- the old nomenclature, GRAD(50 Neg 20), defines the gradients amplitude (50% here), sign (negative here) and its length (in multiplicity of 50msec, means 20X50 msec here). In the present nomenclature, the gradient is defined as a pulse (e.g. p16:gp1), its sign is defined in eda (e.g. GPZ1= -50%) and its shape is defined by gpnam (e.g. GPNAM1= sine.100) .

Pulse Calibration Procedures

renad — Mon, 07 Nov 2022 13:51:24 +0000

Pulse Calibration Procedures renad Mon, 11/07/2022 - 13:51

¹H hard pulse
¹H selective pulse (water Flip Back, FB, pulse)
¹⁵N hard pulse (inverse mode)
¹³C hard pulse (inverse mode)
¹³C selective 90 pulse (shaped pulse)
¹³C selective inversion (180) pulse (shaped pulse)
²H decoupling pulse
Calibrated Pulse-Length Values

¹H hard pulse

Set the power level (pl1) to 0-3 dB (depending on the probe you are using). Type p1 and change the value for small pulse angles to give a small but positive signal (typically start with p1~3-4 µsec). Acquire and process the spectrum (zg, ft, pk) to give a positive signal. [Notice that when the phase correction is defined so that the signal is positive for small pulse angles, then the signal will be positive when the pulse angle is slightly less than 180° and will be negative when the pulse angle is slightly more than 180°]. Change p1, acquire and process the new spectrum (zg, fp), until the signal goes through a null, indicating a 180° pulse.

90deg 180deg

A convenient way to calibrate a 90° pulse is with the automation program paropt. To start the automation program, simply type paropt and answer the following:

Enter parameter to modify: p1
Enter initial parameter value: 16
Enter parameter increment: 0.3
Enter # of experiments: 10

In this case, paropt acquires and processes 10 spectra while incrementing the parameter p1 from 16 µsec to 19 µsec. For each value of p1, only the spectral region defined above (dp1 icon) is plotted. All 10 spectra appear side by side in FILENAME/1/999.
The 1H 90° pulses of the MLEV sequence used during the spinlock period of a TOCSY sequence should be 30 to 40 µsec, while the one required for the cw spinlock used during ROESY corresponds to a 90° pulse length of 100 to 120 µsec. In order to find the corresponding power levels, one can use paropt and modify the parameter pl1 (keeping p1 constant). Alternatively, the user may make use of the rule of thumb that the pulse length should double, approximately, for every 6 dB decrease in power level.

¹H shaped pulse (water flip-back, FB, pulse)

The "water flip-back" is a technique that utilizes field gradients for water suppression by returning the water magnetization to the z-axis prior to acquisition. It uses a selective 90 degree pulse on the water, a shaped flip-back pulse.

On the AVIII800:

Use the 1D pulse program "calib_fb_H.ts", which contains both a 1H 90 degree pulse
{ (p1 ph0): f1 } and the FB pulse { (p2:sp2 ph25:r): f1 }.
First, comment out the FB, run 1D and phase the spectrum.
Comment out the 1H 90 degree pulse and apply the FB. Calibrate a long (p2~ 1-2 msec) 1H shaped pulse (spname2 = sinc1.0) and determine its phase (PHCOR25).
Apply both the 1H 90 hard pulse as well as the selective FB pulse. Optimize the power level of the FB pulse (sp2) so that zero signal will appear. Correct the phase (PHCOR25) of the FB to get the minimal signal.

¹⁵N hard pulse (inverse mode)

The 15N pulse calibration is done in the inverse mode (f1= 1H, f3= 15N), using the pulse sequence DECP90. During calibration, the length and/or strength of the 15N pulse is adjusted. When the 15N pulse is exactly 90°, the anti-phase signal (~5.4 ppm) is minimal. When using the calibration sample 15N-Urea & 13C-Methanol in DMSO, o1p= 5.4 ppm, o2p=76 ppm, p1<90 (~3µsec) and the delay 1/(^2J_XH)= 5.94 msec should be used. The data set UreaMeth 2/1 [in /opt/topspin/data/tali/nmr) is already set to calibrate the 90 hard pulse of 15N. Start with setting the correct phase (to get the anti-phase signal) using a low power 15N pulse, pl3=20 dB (or use a much shorter than desired p21value). Increase the powe level (decrease pl3 values; or, alternatively increase p21) till the anti-phase signal (~5.4 ppm) is minimal.

<90 deg 90 deg >90 deg

¹³Chard pulse (inverse mode)

The 13C pulse calibration is done in the inverse mode (f1= 1H, f2= 13C), using the pulse sequence DECP90. During calibration, the length and/or strength of the 13C pulse is adjusted. When the 13C pulse is exactly 90°, the anti-phase signal (~3.2 ppm) is minimal. When using the calibration sample 15N-Urea & 13C-Methanol in DMSO, o1p= 3.12 ppm, o2p=49 ppm, p1<90 (~3µsec) and the delay 1/(2JXH)= 3.6 msec should be used. The data set UreaMeth 3/1 [in /opt/topspin/data/tali/nmr) is already set to calibrate the 90 hard pulse of 13C. Start with setting the correct phase (to get the anti-phase signal) using a low power 13C pulse, pl2~20 dB (or use a much shorter than desired p3 value). Increase the powe level (decrease pl2 values; or, alternatively increase p3) till the anti-phase signal (~3.12 ppm) is minimal.

Selective Hards Pulses: Off-Resonance Effects

Selective Excitation

In order to avoid excitation of frequencies with an offset dW (Hz), the on-resonance pulse duration for the 90 degree pulse will be
t₉₀ = √15 / 4 (ΔΩ )
where ΔΩ = [basic frequency] * [chem. shift defference in ppm].

Spectrometer Basic Frequencies (MHz):
1H	500.13	800.13
13C	125.757739	201.1927690

Calculated 13C pulse lengths for different spectrometer, to achieve various offsets (ΔΩ), using the above equations:
(o2p)	desired null (ppm)	@ 500MHz	@ 800MHz
		t₉₀(µs)	t₉₀(µs)
43	174	58.7	36.7
53	174	63.6	39.77

Selective Inversion

In order to avoid excitation of frequencies with an offset ΔΩ (Hz), the on-resonance pulse duration for the 180 degree pulse will be

t180 = √3 / 2 (ΔΩ)
and ΔΩ = [basic frequency] * [chem. shift defference in ppm]

Spectrometer Basic Frequencies (MHz)
1H	500.13	800.13
13C	125.757739	201.1927690

Calculated 13C pulse lengths for different spectrometer, to achieve various offsets ( ), using the above equations:
(o2p)	desired null (ppm)	@ 500MHz	@ 800MHz
		t₁₈₀(µs)	t₁₈₀(µs)
43	174	2*26.28	2*16.43
53	174	2*28.4	2*17.79

¹³C selective 90 pulse (shaped pulse)

The selective 13C pulse calibration is done in the inverse mode (f1= 1H, f2= 13C), using the pulse sequence DECP90SP. During calibration, the length (p13 here) and/or strength (sp2 here) of the 13C pulse is adjusted. When the 13C pulse is exactly 90°, the anti-phase signal (~3.2 ppm) is minimal. When using the calibration sample 15N-Urea & 13C-Methanol in DMSO, o1p= 3.12 ppm, o2p=49 ppm, p1<90 (~3µsec) and the delay 1/(^2J_XH)= 3.6 msec should be used. Set the name of the selective pulse (SPNAM2 here) to g4.256. The data set
UreaMeth 4/1 [in /opt/topspin/data/tali/nmr]
is already set to calibrate the 90 selective pulse of 13C. Start with setting the correct phase (to get the anti-phase signal) using a low power 13C pulse, sp2~20 dB (or use a much shorter than desired p13 value). Increase the powe level (decrease sp2 values; or, alternatively increase p13) till the anti-phase signal (~3.18 ppm) is minimal

¹³C selective inversion (180) pulse (shaped pulse)

The selective 13C pulse calibration is done in the inverse mode (f1= 1H, f2= 13C), using the pulse sequence DEC180SP. During calibration, the length (p14 here) and/or strength (sp3 here) of the 13C pulse is adjusted. When the 13C pulse is exactly 180°, the in-phase signal (~3.2 ppm) is minimal. When using the calibration sample 15N-Urea & 13C-Methanol in DMSO, o1p= 3.12 ppm, o2p=49 ppm and the delay 1/(^2J_XH )= 3.6 msec should be used. Set the name of the selective pulse ( SPNAM2 here) to g3.256. Use the calibrated 90 hard pulse values for both 1H (p1) and 13C (p3), long d1 values (4-5 sec), and ns= ds =2. The data set UreaMeth 5/1 [in /opt/topspin/data/tali/nmr) is already set to calibrate the 180 selective pulse of 13C. Start with setting the correct phase (to get the doublet) using a low power 13C pulse, sp3~20 dB (or use a much shorter than desired p14 value). Increase the powe level (decrease sp3 values; or, alternatively increase p14) till the signal (~3.18 ppm) is minimal

²H decoupling pulse (AVIII800)

The use of partially and/or fully 2H labeled samples enables NMR studies of larger proteins in solution. In order to enable alternation of the lock channel between locking and decoupling during the experiment, a free rf-channel and a lock-switch are required. This hardware is available on the AVIII800 spectrometer. The sample used for calibrating the 2H decoupling pulse can be the standard ASTM sample (C₆D₆ in dioxane). The rf routing should enable 13C observation (F1), and 2H should appear on F4. The data set UreaMeth 6/1 [in /opt/topspin/data/tali/nmr) is already set to calibrate the 180 selective pulse of 13C.Use the pulse program decp902hf4, and set o1p=128.3 ppm and o4p=7.28 ppm. The experiment is the same as that used for 15N and 13C inverse 90 calibration (decp90), but is modified to include the 2H switch. At 90 degree, the signal turns into an antiphase that turns its sign when passing the 90.

Calibrated Pulse-Length Values

Calibrated Pulse-Length Values for various nuclei on AVIII800, AV-500, AVIII400 and AVIII300 probes.

CryoProbe: Special Instructions

renad — Mon, 07 Nov 2022 13:07:05 +0000

CryoProbe: Special Instructions renad Mon, 11/07/2022 - 13:07

Introduction
System Overview
Safety & Emergency
Controls on the CryoCooling Unit
Instructions for AVIII800/CryoProbe Users

Introduction

The CryoProbes offer a dramatic increase in S/N by reducing the operating temperature of the NMR coil assembly and the preamplifier. While the sample temperature is stabilized at a user-defined value around room-temperature, the NMR coil assembly -located a few millimeters from the sample- is cooled with cryogenic helium gas.

The most prominent part of the CryoProbes is the cooling unit, in which the so-called ‘coldhead’ expands compressed He and thereby cools it to cryogenic temperatures. Cold He is then circulated through the probe via an insulated He transferline. Vacuum pumps maintain insulation of the probe and the cooler. All operations are supervised by the built-in CryoController unit

System Overview

CryoProbe
Cryo-compatible HPPR
CryoCooling Unit
He cylinder
He Compressor
Water Chiller

Safety & Emergency

The main switch of the CryoCooling Unit serves as an emergency OFF:

powers down the system
all valves are reset to their default positions
the system will slowly warm up

This emergency OFF shuts down the supervisor electronics so it should only be used as a last resort.

The system works with helium gas that is pressurized (25 bar) and cooled to ~20K.

Do not disconnect any tube or cable from a running cold system.
If you have to work with an open cabinet, put on protective goggles and gloves (cold burns)
If large quantity of He gas escapes from the He cylinder during a short period, there is a danger of suffocation and one should care for good ventilation, fresh air supply after the incident
If cold He gas comes in contact with eyes or skin, immediately flood the affected area with cold or tepid water.

Controls on the CryoCooling Unit

As indicated, the main switch on the CryoCooling Unit front serves as an EMERGENCY OFF.
In addition to the main switch on the CryoCooling Unit front, six additional buttons exist:

WARM
WARM UP
COOL DOWN
COLD
ERROR
UNPLUG

All buttons and indicators may illuminate to give a status display
WARM UP, COOL DOWN and UNPLUG buttons can be pressed for input.
When COLD lights up (green), the system is ready for measurements
When COLD blinks, the system is slightly overheated- check and reduce decoupling !!
Whenever other buttons blinks/lights up, pls call immediately an authorized person

Instructions for AVIII800/CryoProbe Users

Choose the appropriate probe-head, using the edhead command (probe #38)
Due to the sensitivity of the system to the temperature at the sample cavity, load the configuration for the VT unit via edte: edte -> load configuration (choose a configuration file that best matchs your working conditions in terms of probehead (TCI), temperature, flow-rate, BCU unit status -ON or OFF- state etc).
Use the white (preferably) or blue spinner (never use the ceramic one!!)
Avoid fast dropping of samples with the sample lift.
Do not introduce any objects into the sample cavity of the probe (not even soft cotton bud)
Insert the sample only if the COLD button on the cooling unit is ON, or at least flashing
The gas flow must be maintained at all times while the CryoProbe is cold- it must not be interrupted before the CryoProbe has been warmed-up
The sample is subjected to a rather high flow of VT gas (800-935 l/hr).
Set the max VT heater power limit to 20% (as low as possible)
Use temperature in the range of 0C to +60C only
Always keep an eye on the sample temperature (edte)- if the temperature drops, eject the sample to prevent freezing. Never interrupt the VT gas flow !!
CryoProbe must not be tuned or matched when warm- before attempting to wobb the CryoProbe, the green COLD light should be ON or at least flashing
Tuning & Matching the CRP:

An Automatic Tuning and Matching (ATM) unit was recently installed.
The wobbling procedure could be done now either automatically using the atma command, or manually (by clicking the appropriate bottons on the displayed ATM window) using the atmm command.
PLS NOTE: For salty samples (>100mM salt) pls use the manual option, atmm.
The old wobb command could only be used to view the wobbling signals (no changes could made from that window).

Verify (via edasp ) that the RF routing in your file is in accordance with the new HPPR-CRP rules

frequency      amplifier        preamplifier
F1   1H       FCU1     H 100.0W      1H
F2   13C     FCU2     X 500.0W      13C
F3   15N     FCU3     X 300.0W      15N
F4   2H       FCU4     X 300.0W   locksw 2H 2H

Excessive RF power can destroy the CryoProbe or its HPPR- obey the limitations. If in doubt, start with powers that are at least 10 dB below (more positive values) the values you are used to.
In any case, follow Bruker application-specialist Instructions , found at the folder near the magnet.
Calibrated pulse lengths values

1H         10.12 us @ +5.0dB
13C      12.17 us @ -1.50 dB
15N      39.48 us @    -0.80 dB

When 2 RF hard pulses are applied to a NMR coil at the same time (e.g. pulsing 13C & 15N), reduce the power of the two pulses by 3 dB each.
In case of 3 simultaneous hard pulses, the reduction is 10 dB for each.
The decoupling power required by the CryoProbe is much smaller than a conventional probe- in case the decoupling cases heating of the probe, the COLD button on the cooling unit will start flashing -should this occur, stop immediately the experiment, reduce the decoupling power, and call an authorized person. Do not continue the experiment.
Since no attenuators are connected to the 3 amplifiers at the console, a special care should be taken regarding excessive pulsing power!!
A characteristic periodic hiss of the cooling unit is clearly audible at all times
In normal measuring conditions, the COLD green button lights permanently. In case of blinking, or if other buttons lights/blinks, pls call immediately authorized people.
All necessary filters are already incorporated in the system- no additional filters are required
If a large quantity of He gas escapes from the He cylinder during a short period, there is a danger of suffocation- take care for a good ventilation and fresh air supply after the accident
If cold He gas comes in contact with eyes or skin, immediately flood the affected area with cold or tepid water.

Sample-Temperature Calibration

renad — Mon, 07 Nov 2022 12:59:13 +0000

Sample-Temperature Calibration renad Mon, 11/07/2022 - 12:59

Molecular Associations can lead to shifts in the NMR spectra. This effect is particularly strong, when hydrogen bonding is involved. The shifts are concentration dependent and temperature dependent. This fact is used when calibrating the actual temperatures in your sample with methanol (low temperature range, 178-330K) and/or ethylen glycol (high temperature range, 273-416K).

We have used neat methanol as our thermometer substance, and measured the frequency difference between its two peaks to calculate the actual sample temperature.

Temperature calibration for AVIII-800 probes

Bruker TCI CryoProbe 5mm (#38):
Sample Set/target Temp.	Actual Sample Temp.
312 K	311.3 K
307 K	306.4 K
302 K	301.5 K
297 K	296.4 K
292 K	291.7 K
288.9 K	288.7 K
287 K	286.8 K
282 K	281.9 K
277 K	277.1 K

Bruker QXI 5mm probe (#37):
Sample Set Temp.	Actual Sample Temp.
308 K	K
303 K	K
295 K	K
290 K	K
285 K	K
280 K	K
277 K	K
275 K	K

Temperature calibration for NEO-600 probes

Bruker TCI CryoProbe 5mm (Z159838_001):
Sample Set/target Temp.	Actual Sample Temp.
312 K	K
307 K	K
303 K	303 K
298 K	298 K
293 K	293 K
288.9 K	K
287 K	K
282 K	K
277 K	K

Bruker TXI 5mm probe (Z816801_0154):
Sample Set Temp.	Actual Sample Temp.
308 K	K
303 K	K
298 K	K
293 K	K

Calibrated Pulse-Length Values

renad — Mon, 07 Nov 2022 12:41:30 +0000

Calibrated Pulse-Length Values renad Mon, 11/07/2022 - 12:41

Calibrated pulse lengths for NEO-1000 probes
Calibrated pulse lengths for AVIII-800 probes
Calibrated pulse lengths for NEO-600 probes
Calibrated pulse lengths for AV-500 probes
Calibrated pulse lengths for AVIII-400 probes
Calibrated pulse lengths for AVIII-300 probes
Pulse Calibration Procedures

Calibrated pulse lengths for NEO-1000 probes

Probe	Nucl	90 deg Pulse length (µs)	Power level (dB)	Remarks

RT-5-TXI	1H	10.0	-13.25	hard
	13C	13.00	-18.59	hard
	"	25	-18.09	C-C tocsy
	"	50	-12.07	dec, garp
	"	246	-23.65	90deg sp, g4.256
	"	900	-12.23	90deg sp, Q5.1000
	"	154	-24.71	180deg sp, Q3.1000
	"	630	-14.46	180deg sp, Q3.surbop.1
	"	500	-17.89	Crp60 ; Adiabatic inversion
	"	2000	-17.89	Crp60comp.4; Adiabatic refocusing
	15N	39.0	-23.77	hard
	"	220	-8.28	cpd, garp
	2H	150	-18.59	hard
		176	-17.21	dec

CP 5-TCI-X,Y,Z	1H			hard
	"			tocsy*
	13C	12		hard
	"	13.14		offres=131ppm
	"	14.23		offres=121ppm
	"	25		C-C tocsy
	"	29.39		offres=131ppm
	"	31.82		offres=121ppm
	"			garp dec
	"
	"	333		sp, g4
	"	512		sp, g4
	"	256		sp, g3
	"	512		sp, g3
	"			sp, g3 (sel CO dec)
	15N			hard
	"	220		dec, garp
	2H

CP 3-TCI	1H			hard*
	13C			hard
	"	25		C-C tocsy
	"			garp dec 100ppm
	"
	"	333		sp, g4
	"	512		sp, g4
	"	256		sp, g3
	"	512		sp, g3
	"			sp, g3
	15N			hard
	"	220		dec, garp
	2H
	2H

Calibrated pulse lengths for AVIII-800 probes

Probe	Nucl	90 deg Pulse length (µs)	Power level (dB)	Remarks

CRP-TCI (#38)	1H	14.5	-8.05	hard*
	13C	14.35	-21.06	hard
	"	16.43		offres=131ppm
	"	17.64		offres=122ppm
	"	25	-16.5	C-C tocsy
	"	36.7		offres=131ppm
	"	39.45		offres=122ppm
	"	67	-8.04	dec, garp
	"	333	-19.78	90deg sp, g4.256
	"	333	-18.95	90deg sp, Q5.1000
	"	410	-17.33	90deg sp, Q5.1000
	"	512	-16.00	90deg sp, g4.256
	"	256	-19.15	180deg sp, g3.256
	"	256	-18.60	180deg sp, Q3.1000
	"	310	-16.97	180deg sp, Q3.1000
	"	512	-12.62	180deg sp, Q3.256
	"	620	-10.92	180deg sp, Q3.1000
	"	256	-6.90	sp, sinc.128
	"	256	-8.87	sp, Seduce.100
	15N	47.71	-20.87	hard
	"	190	-8.59	dec, garp
	2H	184	8.2 W	hard
		244.5	4.4978 W	dec
		433.5	1.5 W	dec

TXI	1H	10.2		hard*
	"	30		tocsy*
	"	100		roesy*
	"	1000		selective (H2O)
	13C			hard
	"	16.3		offres=132ppm
	"	17.64		offres=122ppm
	"	25		C-C tocsy
	"	36.46		offres=132ppm
	"	39.45		offres=122ppm
	"	60		garp dec 100ppm
	"	67
	"	333		sp, g4
	"	512		sp, g4
	"	256		sp, g3
	"	512		sp, g3
	"	768		sp, g3 (sel CO dec)
	15N			hard
	"	210		dec, garp
	2H	300


QXI	1H			hard*
	13C			hard
	"	16.3		offres=132ppm
	"	17.64		offres=122ppm
	"	25		C-C tocsy
	"	36.46		offres=132ppm
	"	39.45		offres=122ppm
	"	60		garp dec 100ppm
	"	67
	"	333		sp, g4
	"	512		sp, g4
	"	256		sp, g3
	"	512		sp, g3
	"	768		sp, g3
	15N			hard
	"	210		dec, garp
	2H	300
	2H	500

*Depends very much on the sample. Should be calibrated again on the studied sample.

Calibrated pulse lengths for NEO-600 probes

Probe	Nucl	90 deg Pulse length (µs)	Power level (dB)	Remarks

5 mm CRP-TCI	1H		-8.32	hard*
	13C	11.8	-20.49	hard
	"	27	-13.18	C-C tocsy
	"	67	-5.12	dec, garp
	"	210
	"	333	-16.47	90deg sp, Q5.1000
	"	400	-14.86	90deg sp, Q5.1000
	"	256	-15.75	180deg sp, Q3.1000
	"	310	-14.03	180deg sp, Q3.1000
	"	512		180deg sp, Q3.256
	"	620		180deg sp, Q3.1000
	"	128	-10.0	sp, sinc.128
	"	256		sp, Seduce.100
	15N	36.5	-21.02	hard
	"	170	-7.48	dec, garp
	2H	76.3	-15.8	hard
		463	-0.14	dec

1.7 mm CRP TCI	1H	10.2		hard*
	13C			hard
	"	25		C-C tocsy
	"	67
	15N			hard
	"	210		dec, garp

5 mm TXI	1H			hard*
	13C			hard
	"	25		C-C tocsy
	"	60		garp dec 100ppm
	"	67
	15N			hard
	"	210		dec, garp

*Depends very much on the sample. Should be calibrated again on the studied sample.

Calibrated pulse lengths for AV-500 probes

PROBE	Nucl	High Power		Low power
		p1 (µs)	pl1 (dB)	p1 (µs)	pl1(dB)

BBI	1H	6.8	0
	13C	13	-2	67	12
	15N	27	-2	208	16
	31P	17.8	0	72	12

BBO	1H	7.5	-4	110	19
	13C	7	-1
	31P	7.6	-2
	15N	10	-4

TBI	1H	7.3	0
	13C	13.2	-3	69	11
	15N	21.5	-4	188	15
	31P	20.5	-1	72	10

TXI	1H	3.6	0
	13C	9.6	-4	70	13
	15N	27	-4	195	13
QNP	1H	11	-6	100	14
	13C	6	-6
	31P	8.2	0	100	21
	19F	10.2	-6

Calibrated pulse lengths for AVIII-400 probes

PROBE	Nucl	High Power		Low power		remarks
		p1 (µs)	pl1 (dB)	p1 (µs)	pl1(dB)

QNP	1H	13	-4	80	14
	13C	6.2-5.8	-6	80	19
	31P	6.5	-2	80(90)	15(26)
	19F	15	-6	80	10

BB5	1H	9	-4	80	19
	13C	17	-4
		14.4	-6
	2H	11	-4
	15N	12.6	-4
	17O	14	-4
	29Si	9	-4
	31P
	51V	7.2	-4
	119Sn	6.15	-4			no dec.
		5.75	-4			with dec.
	121Sb	8.8	-4
	127I	12.5	-4
	195Pt	12	-4

	13C	18	20W
	19F	14	12W
	23Na	11	50W


BBI	1H	6.7	-3	80	18	ROESY spin-lock 22dB
	13C	13	-6	85	12	100W
		9.6	-3	90	17	300W
				85	16	300W
	15N	26.5	-4	180	13	100W
				200	14	100W
		16	-4	180	18	300W
				200	19	300W
	31P	12	-4
	29Si	15	-6	130	12	100W
		11.4	-3	80	14	300W

TBI	1H	6.5	-3	80	17	100W
	13C	9	-4			300w
	15N	26	-4			100W
		14(16)	-4	190	19	300W
	31P	23.5	-4			100W

BB10	1H
	17O	28	-4
	31P	22	-4	100	10

SEL	1H	7.9	-3

BBO10	39K	38	-6
	183W	40	-6

BBFO	1H	13	18.5W	90	0.411W
	2H	30	25W
	11B	16	30W
	13C	9	64W	70	1.2W
	14N	15	60W
	15N	19.5	66.5W	180	0.85W
	19F	13	30W	70	1.0347W
	29Si	11.5	65W	180	0.25W
	31P	13	17W	80	0.45W
	31P	20	26W	80	1.625W
	33S	30	60W
	77Se	12	60W

Calibrated pulse lengths for AVIII-300 probes

PROBE	Nucl	High Power		Low Power
		p1 (µs)	pl1 (dB)	p1 (µs)	pl1 (dB)

QNP	1H	11	-6	100	14
	13C	6	-6
	31P	8.2	0	100	21
	19F	10.2	-6

BB5	1H	10.5	-6	100	17
	13C	5.5	-3
	31P	6.7	0

Pulse Calibration Procedures

Procedures for calibrating different pulses for various nuclei are described.

Shimming

renad — Mon, 07 Nov 2022 12:26:47 +0000

Shimming renad Mon, 11/07/2022 - 12:26

In the context of NMR, the shims are small magnetic fields used to cancel out errors in the static magnetic field. These minor spatial inhomogeneities in the magnetic field could be caused by the magnet design, materials in the probe, variations in the thickness of the sample tube, sample permeability, and ferromagnetic materials around the magnet. A shim coil is thus designed to create a small magnetic field which will oppose and cancel out an inhomogeneity in the B o magnetic field. The most fundamental criterion of the homogeneity is the observed line shape, and in modern spectrometers, the two measures of homogeneity is the deuterium lock signal and the free induction decay (FID).

The deuterium lock is the mean by which long term stability of the magnetic field is achieved. The field strength might vary over time due to aging of the magnet, movement of metal objects near the magnet, and temperature fluctuations. The field lock can compensate for these variations. The field lock is a separate NMR spectrometer within the spectrometer. This spectrometer is typically tuned to the deuterium NMR resonance frequency. It constantly monitors the resonance frequency of the deuterium signal and makes minor changes in the Bo magnetic field to keep the resonance frequency constant.

The lock level is a convenient indicator of the homogeneity: the lock signal results from observing a single line, usually of the deuterated solvent. The area under this line is constant, but the width of the line depends on the homogeneity: as the line becomes narrower it must also become taller, thus the lock signal, displayed as the lock level, is higher. Thus the object is to maximize it

Adjusting the shim

There are different shim coils that can create a variety of opposing field. By passing the appropriate amount of current through each coil a homogeneous magnetic field can be achieved. The optimum shim current settings are found by either maximizing the signal from the field lock or maximizing the size of the FID. While shimming, our task is to find the best shim value by maximizing the lock signal.
One should take into consideration that high order shim fields (e.g. Z⁴) are 'contaminated' with shim fields of lower orders with the same parity (i.e. Z³ contains contributions from Z; Z 4 from Z²). So when adjusting high order components (Z³ , Z⁴ etc) it is essential to re-optimize the lower order components (Z, Z² here).
A shim protocol starts with adjusting
Z, Z²
and then the low order horizontal gradients
X, XZ
Y, YZ
XY, X² -Y²
and again
Z, Z² .
Than go to higher orders if necessary
Z³ , Z
Z⁴ , Z²
Z⁴ , Z³, Z.
Try to improve the horizontals, and the low Z, again.

The shape of an NMR line is a good indication of which shim is misadjusted:

(taken from Modern NMR Techniques, by Andrew Derome, 1987)

Shimming using the FID

If the FID can be displayed in real time (using the Bruker command 'gs'), the magnetic field can be shimmed by observing the shape of the decay envelope. Two features of the appearance of the FID are informative: its duration and its shape. The duration of the decay gives information about the eventual linewidth, and the appearance of the decay envelope gives information about its line shape:
(A) correct FID shape (B) Incorrect FID shape

(taken from Modern NMR Techniques , by Andrew Derome, 1987)

The simple exponential shown in (A) above, is only observed with samples whose signal is dominated by a strong single resonance line (e.g. by that of the water solvent).

Gradient Shimming

The old gradshim feature, automatic shimming using gradients, is available on probes that are equipped with gradients. It requires the existance of reference mapping (both 1D as well as 3D maps) for each probe.

TOPSHIM

The topshim feature, a completely automatic shimming using gradients, is available on the AVIII800, NEO600, AVIII400 systems. For its graphic interface use the command topshim gui.

Nice tips on shimming

Solvent Suppression

renad — Mon, 07 Nov 2022 12:22:11 +0000

Solvent Suppression renad Mon, 11/07/2022 - 12:22

Detection of the solute signal (typically proton concentration of 1-2 mM) in the presence of solvent signal (for water, proton concentration ~110M) presents a difficult problem since the dynamic range of the electronic components of the spectrometer is limited.
Three methods exist for suppressing unwanted signals of the solvent (dynamic range reduction):

Presaturation

of the solvent resonance during the recycle delay between the acquired scans, using a weak rf field. Presaturation of a signal excites a small region of the sample for a relatively long time, reducing the intensities of the net magnetization of signals in the region. The main disadvantages are that signals that resonate very close to the solvent may be partially saturated (the alpha protons of the protein). Also, if nuclei are exchanging between two environments that give rise to two different signals, and if one of these signals is presaturated and reduced in intensity, then the other signal will also be reduced (saturation transfer). For instance, if presaturation is used to reduce the water signal in a protein sample, the amide proton signals will also be reduced, since the amide protons exchange with the water proton. In order to use this water suppression scheme, set the pulse program to zgpr, set your carrier frequency o1 on the solvent, and irradiate for d1 of ~1-2 sec using an rf field of ~50 Hz (typically 58dB for H2O sample, and 80-100dB for samples in D₂O).

Binomial signal suppression

experiments use non-uniform excitation of the spectrum to reduce the signal intensities of small spectral regions. The 1-1 jump-return (Plateau & Gueron, JACS104, 1982) and the 1331 binomial sequence (Hore, J. Mag. Res. 55, 1983) are recommended for 1D signal suppressions. The higher-order binomial experiments better suppress peaks at the expense of a more rolling baseline. Suppression will occur at multiplets of the offset frequency. In the jump-return technique, the final read pulse in a pulse program is replaced by the sequence 90- t -90, the carrier is placed on the solvent, and the delay is set to t = 1/(2* d ) where d is the offset from the carrier to the next null. In the 1331 sequence, p1331, the delay d19 = 1/d where d is the offset from the carrier to the next null. The left half and right half of the spectrum differ in phase by 180°, and the spectrum may require lots of first-order phase correction.

Dephasing of the solvent

using spin lock and gradient pulses. There are many sequences that uses these elements to achieve solvent suppression: some uses spin-lock purge pulse in which the solute magnetization is locked while the solvent’s dephase; some use strong field gradients to dephase the solvent (WATERGATE; Piotto, Saudek & Sklenar, J. Biomol. NMR 2, 1992) while others return the water magnetization to the z-axis prior to acquisition (flip-back; Grzesiek & Bax, JACS115 , 1993). In the WATERGATE sequence, zggpwg , the gradient pulse is typically 1msec and shaped selective pulses are used. An easily set-up sequence that dephases the solvent by combining the binomial water suppression with gradient pulses is the p3919gp pulse program. It requires no special calibrations since it uses solely hard pulses and gradient pulses. As in the 1-1 jump-return sequence, d19=1/2*d where d is the offset from the carrier frequency, o1, to the next null.

NMR

AVANCE NEO-400 WB (Bruker)

Short Spectrometer Manuals

The Linux/UNIX operating system

Interacting with the TopSpin program

Data Acquisition

Defining data sets

Reading and editing parameters

Setting the temperature

Important acquisition parameters

Locking and Shimming

Tuning the probe

Calibrating the pulses

Acquiring the data

Important note

Data Processing

Other Useful Tasks

A brief summary of the steps

Pulse Program Information and Properties

The two-character codes used are the following

Common Bruker Commands and Nomenclature

Pulse Calibration Procedures

1H hard pulse

1H shaped pulse (water flip-back, FB, pulse)

15N hard pulse (inverse mode)

13C hard pulse (inverse mode)

Selective Hards Pulses: Off-Resonance Effects

Selective Excitation

Selective Inversion

13C selective 90 pulse (shaped pulse)

13C selective inversion (180) pulse (shaped pulse)

2H decoupling pulse (AVIII800)

Calibrated Pulse-Length Values

CryoProbe: Special Instructions

Introduction

System Overview

Safety & Emergency

Controls on the CryoCooling Unit

Instructions for AVIII800/CryoProbe Users

Sample-Temperature Calibration

Temperature calibration for AVIII-800 probes

Temperature calibration for NEO-600 probes

Calibrated Pulse-Length Values

Calibrated pulse lengths for NEO-1000 probes

Calibrated pulse lengths for AVIII-800 probes

Calibrated pulse lengths for NEO-600 probes

Calibrated pulse lengths for AV-500 probes

Calibrated pulse lengths for AVIII-400 probes

Calibrated pulse lengths for AVIII-300 probes

Pulse Calibration Procedures

Shimming

Adjusting the shim

Shimming using the FID

Gradient Shimming

TOPSHIM

Solvent Suppression

Presaturation

Binomial signal suppression

Dephasing of the solvent

¹H hard pulse

¹H shaped pulse (water flip-back, FB, pulse)

¹⁵N hard pulse (inverse mode)

¹³Chard pulse (inverse mode)

¹³C selective 90 pulse (shaped pulse)

¹³C selective inversion (180) pulse (shaped pulse)

²H decoupling pulse (AVIII800)