Transcript Introduction to ICON NMR This slide presentation is a brief introduction to the use of the data acquisition software ICONNMR.

Introduction to ICON NMR
This slide presentation is a brief
introduction to the use of the data
acquisition software ICONNMR. It is
not intended to take the place of formal
hands-on instruction.
Revision date 12/02/04
Important information. We have now fixed ICON so that
spectra will be automatically emailed to your email account.
See the new info starting on approx. slide 80.
Revision 12/27/04
Two bugs and their fixes are described at the end of the
presentation.
REVIEW OF THE MAJOR NMR LAB
RULES
•
•
•
•
No food, and of course no smoking.
No drinks of any kind, including water.
No loose metal objects, especially paper clips and staples.
No chemistry at or near the consoles. A wet prep lab is available for
adding things to your nmr tube.
• Remember: Mechanical watches and credit/ID card magnetic strips
are erased by the magnetic field.
• DO NOT walk near any magnet except the one you are using.
• Be quiet and courteous of others working in the lab.
The basic operating system of the acquisition computer and
the lab data station is RedHat Linux. This is a multi-user
OS similar to what you see on bama.ua.edu. Every nmractive research group has its own account listed by the
name of the principle investigator. Every major course also
has an account listed by the course number. Within these
accounts, ICON provides for the creation of specific nmr
users. NMR administration (Admin) will provide you with
the user name and password for your research group
account (i.e., account timkovich, password ********). Do not
share this information with people outside your group, or
they will have access to your data files. Do not change the
password. Admin will need it to help administer your
account.
The group leader will provide Admin with a list of
group nmr users (grad students, postdocs, etc.).
Each user will be assigned an ICON user name
and password. Only Admin can issue these. It is
recommended that you keep your ICON password
confidential, even from other group members.
ICON has tracking capabilities, and if you want to
be blamed for someone else’s mistakes, give them
your ICON password. Your data will be stored in a
directory unique to each ICON user, but, everyone
in your group will have access to the directory.
People outside your group will not.
The following slides are mainly screen captures
that have been edited with comments that show the
major windows that open during ICON. Not all
possible windows can be shown. Ignore the small
boxes that have import commands in them; they
were used to capture the screen.
Type your group account name, hit enter, then type
your group password and hit enter.
Click here to start
Main program called Topspin is
starting
Group home directory
user directory
data file
Click here
Move cursor here
Click here
Click here
Then click OK
Click on your
username
This box is too small
Place cursor on right edge
drag to increase size
Enter your ICON
password, then hit
enter
This box will be flashing red
Click on it
This new window opens. It is also too small.
Place cursor on right edge and drag to
enlarge.
Click here and wait. The sample inside
the spectrometer will slowly rise to the
top.
A new window appears. Leave
it open while you load your
sample tube into the spinner.
Use the stepladder. Do not touch or lean on the magnet. Remove
the spinner from the magnet top with the previous sample which
should be a standard sample of D2O. Remove the standard from
the spinner and place it in the empty box on the top of the console.
There should always be a sample in the magnet even in between
In order to keep dust out of the probe and the spinner turbine.
Instructions for sample preparation
Nowadays, the spectrometers use a fairly sophisticated
automated routine for sample shimming: adjusting the magnetic
field homogeneity to produce the best resolution, lineshape, and
peak height. This eliminates the tedious and time consuming
manual adjustment of shims, that was also highly dependent on
the individual operator. However, for the automated routine to
be successful, you must adhere to these guidelines for sample
preparation.
Use only approved tubes as on the next slide.
Use the correct deuterated solvent for your sample. The
presence of TMS is optional, but if used do not use too much.
In general, 1% TMS is an overkill for modern instruments.
TMS is not really needed for referencing. The residual protic
solvent in your deuterated solvent is wholly adequate.
No precipitate or solids in the sample. These kill shimming and
potentially tuning. Filter as needed.
Sample Prep, cont.
Use a syringe and fill the tube to an accurate volume of 0.800 mL.
Automated shimming starts with standard shim files that have
been created for the probe. These were all based upon
standard samples containing this volume. The success of
automated shimming depends critically on the correct
positioning of the sample with the depth gauge (to be discussed) and
on the sample volume. 0.8 mL corresponds to a liquid depth of
58 mm. A little bit more volume is better than ANY less.
Instructions for NMR tubes.
In high field instruments, the sample coil fits VERY tightly around the
spinning NMR tube. If the tube is dirty on the outside, material is
transferred to the probe coil. This can degrade the probe tuning, scratch
the coil, ruin the shimming, or create unwanted and unremovable
background signals to spectra. So:
Rule 1. CLEAN the outside of your sample tube with acetone and wipe
it dry BEFORE coming to the NMR lab.
The quality and cost of tubes relates to several factors. Most important to
the safety of the probe is the straightness of the tube and its ability to
spin without wobble. So:
Rule 2. Use only Wilmad 528 or Kontes 235 tubes or tubes of higher
quality.
Unfortunately, there are a variety of other cheaper,inferior tubes still
present in the Department from older days, etc.. Probes cost
$40,000. The cost difference between tubes is a couple bucks. We
cannot afford a mistake. So:
Rule 3. The tube MUST display the glass etcing mark of “528” or
“235” or higher numbers.
Rule 4: NEVER use a tube that is chipped or cracked at the top.
These are weak and can break in the probe.
Rule 5. Do NOT have the glass shop cut off the tops of damaged
tubes so that the etch mark is gone.
Rule 6. If Admin catches a non-approved tube, it will be
confiscated and DESTROYED, regardless of contents.
The following photo's demonstrate how to insert the tube into the
spinner, use the depth gauge to position the tube properly in the
spinner, how to wipe off the tube and spinner with a Kimwipe
(IMPORTANT), how to hold the tube and insert it into the
magnet.
CRITICAL!!!! THE COLUMN OF AIR THAT EJECTED THE
PREVIOUS SMAPLE MUST STILL BE FLOWING. YOU
MUST HEAR THE AIR FLOW!!!
CRITICAL!!! DO NOT LEAN ON THE MAGNET OR PUT YOUR
HANDS ON THE MAGNET. USE THE STEPLADDER
PROVIDED.
1. Capped and clean NMR tube. 2. Spinner. We have several kinds:
blue and white plastic and a special ceramic spinner for high
temperature work. 3. Depth gauge. This is a precision tool. Do not
play with it or change any of its settings. Leave it on the console
top. Never move it near the magnet.
3. Depth gauge
1. Standard D2O
sample
2. Spinner
Clean the tube!
Wipe the bottom of the
spinner
Keep fingerprints off!!!!!
Gently push tube until tube bottom touches white disk inside
gauge. Do not play with the gauge! It is a precison tool. Do
not change its settings!!!!!
The gauge goes back into its
box.
NOT near the magnet.
NOT on the computer table.
DON'T approach with credit cards.
DON'T appraoch with your wallet.
DON'T approach with a watch on.
DON'T touch the magnet.
DON'T lean on the magnet.
DO use the stepladder provided.
Proper way to grasp the
spinner and sample.
Keep fingerprints OFF!
Gently insert spinner. YOU MUST HEAR AIR
FLOW!!!!
Use the ladder.
Hands off the magnet!!
With your sample in position
click
OK.
After the sample loads a new window
appears.
Explained on next slide
After noting information, click OK
A swept signal showing your
deuterium solvent.
Disk: /home/belmore This is the directory where your data will be
stored. “belmore” will be your group account.
File: kab-111504
No: 3
This is a directory name, not really a file.
Your spectrum will be stored in this directory.
The name is assigned automatically. “kab” is
your ICON user name. “111504” is the date
in the format “month-date-year”
This is called the exp number. It is assigned
automatically. The first time you use ICON
on this day it will be set to 1. Thereafter,
every time you use or enter ICON, it will be
incremented by 1.
Now we have to explain how Bruker actually stores data. The
first issue is to clarify a major difference between the new
systems and the old ones. In the old system, data collected
resided only in computer memory until you specifically saved it
by writing a file. In the new system, the hard disk space
is reserved BEFORE any data is collected. After an acquisition,
but not during it, the data is written from the slave acquisition
computer to the hard disk.
Data is stored in a complicated directory structure. “File name”
is actually this directory name. It contains not only your fid, but
boatloads of other files that contain all sorts of information on
how the fid was acquired, the current status of the instrument,
etc.. Just about everything short of what you had for
lunch today. This is a consequence of the fact that hard disk
storage has become so cheap, and nmr data, in contrast to audio
files, dvd's, etc., is comparatively small in size. Rather than risk
missing anything, Bruker packs everything into this directory.
The first raw fid that you acquire will be placed into a subdirectory
labelled “1” that is actually the “exp” subdirectory. ICON will
automatically process this fid into a spectrum and store it in a subsub directory to “1” called “proc” with the label “1”. If an expert
user re-processes fid“1” say with different line broadening, then the
new spectrum will be saved as proc “2” within the exp “1”
subdirectory. Reprocessing again leads to proc “3” and so forth.
The next fid that you collect on this day will be stored automatically
by ICON into the same master directory ”file name” (for example,
kab-111504) but in an exp subdirectory labelled “2”. “2” will have
processed spectra“1” and higher if it has been reprocessed.
Then the next fid will be exp “3” and so on. It is up to you to
keep track of what you collected in exp “3” and so forth.
However, we will give you an opportunity to include your own
descriptive information in a title.
The next slides illustrate where TOPSPIN displays the exp and
proc numbers.
Master directory kab-111204
exp no. 1, i.e. First fid
proc no., i.e. First processing
4th proc of 10th fid
After noting information, click OK
A swept signal showing your
deuterium solvent.
Back to this window. This box will
be blinking red. Click on it.
In this window, click on this down
arrow icon. A menu appears with a list
of deuterated solvents. Click on the
solvent for this sample.
Note that the solvent name
appears in the box. Make
sure it is correct!
Click on this icon. Another
menu appears with a list
of experiments that you are
allowed to perform. Select
the proper experiment.
Allowed experiments
This list is under constant development and is changing daily.
Admin will post a list separately on the bama_nmr web site.
The two most frequently used experiments are UAbasicH1
(standard routine proton) and UAbasicC13 (standard carbon
with proton decoupling).
Do not select any other experiment until you have been
trained in its use. You can damage the spectrometer by
attempting to run the wrong experiment.
Note proper experiment
has been selected.
If you desire to add a title
or other information for
description, click here
Another window will open entitled Set Title.
Type any descriptive text you wish and then
click on the Set button. The beginning few
characters of your message (but not the entire
message) appears here.
At this time you have the option of modifying
some of the basic default settings for your
experiment. If you wish to do so click here.
A new menu appears, click on the item
User Specific Commands
User Specific Commands
These allow you to change certain standard defaults. When you
click on this, still another menu appears. Common options include
sw
ds
si
sweep width in ppm of the spectrum.
discard scans, number of dummy scans before real data.
size. The number of digital data points in the spectrum.
The bigger, the more digital resolution, but note that almost all
spectra are limited by the magnetic resolution. The default
is normally very adequate.
O1 The center of your spectrum in Hz.
ns number of scans. Decrease to save time, increase to
improve signal averaging.
expt experiment time. Clicking on this will show the total time to
collect all data. This does not include setup time, e.g., shimming.
This list will be continually updated.
Click on the user command you wish to modify. A new dialog
box appears showing the current value and allowing you to
type in another value. Click the OK button when finished.
Without further training do not alter any other commands.
After accepting the defaults or
making changes, click here.
This icon will be flashing red.
At this point you can go back
to any of the previous menu's
and make changes. Click on
this icon to start data collection.
This icon will be blinking
This new window appears
Then this icon will start
blinking
Then this icon starts blinking.
It will continue for 4-8 minutes
while it does auto-shimming.
If you have your windows sized
right, you can follow status
messages here during autoshim.
Auto-shimming
Depends on probe and sample. The protocol is programmed by
Admin. The current protocol is a compromise between the
quality of the final spectrum and the time spent shimming. The
current protocol produces quality results if you have followed
the previous sample prep instructions.
Do not be alarmed
by this message.
The shims will still
be OK
This blinks
for a few
seconds
This blinks while acquisition
is in progress.
You can follow the time
remaining here
This blinks once or twice, then
finishes
Now exit ICON by clicking
File,
then Exit.
Your
title
You are now in the Topspin
acquisition window. This is
not needed. Click here to
close.
This window appears containing
your automatically processed
spectrum. You only need to
check it for spectral quality.
To increase vertical scale click
here.
To expand horizontally, place
the cursor inside the window.
Right click and hold down the
button. Drag the yellow line to
the opposite border of the
expansion. Release the button
and an expanded spectrum
appears.
If you did not get what you wanted,
click here and the full spectrum will
appear. Then repeat expanding as
needed.
Notes on Processing
The auto-processing routine will apodize your fid, Fourier
transform it, phase correct it, baseline correct, and assign an
APPROXIMATE chemical shift scale. This will allow you to
determine whether the spectrum was successfully recorded.
Further processing is to be done by you offline, using any
processing package you wish, such as MESTREC, Swan, Acorn,
etc.. You are NOT to integrate or plot on the spectrometer.
If you are happy with the quality of the spectrum, you will follow
the steps to email the spectrum to a valid email account.
But let's say you need to tweek the results. You want to collect
more scans and change the sweep width.
Restart ICON Main, then Routine. Log back in. Click on the
Inject icon. The following window appears.
Click here. Icon will collect
another fid, but will skip locking
and shimming and save time.
ICON re-opens this
window.
Re-set your solvent and
experiment
Now click OK
This is blinking red. Click here to go
to the filename page and autoincrement the exp number. Note
that on this restart, ICON jumped
from Inject to Set solvent, then to
Filename, whereas on a first start,
it opens the windows in order.
Go figure. Regardless, all info must
be updated before a fid can be
collected.
The exp number will be autoincremented. Make note of it
for your records, then click
OK
This will be blinking red awaiting the
click to start acquisition. But wait!
The purpose of this restart was to
change exp conditions, such as ns.
Click on parameters, then User
Specific, then the parameter to be
changed and enter the new value. Then
Start
Explanation
Certain ICON windows that open, like the previous Parameter
window, are active windows that expect an immediate action.
The screen capture command called “import” that is being
used to generate this tutorial does not allow one to capture
an active window without first closing it. Hence active windows
cannot be displayed in this presentation.
The top icons blink in order.
Locking and shimming are
skipped since you told it to Use
Same. Now File and Exit to
leave ICON.
For long experiment (e.g. C13) Users
Use the Parameters>User Specific>ns sequence to set ns to
some number, like 500. Then use the expt command to
calculate the time for this ns. Calculate how many ns are
needed for your block of time so the exp will finish at the end.
We will no longer use the ns= -1 command. It is your
responsibility to come back at the end of your block of time,
email your results to an offline acount, remove your sample and
re-insert the standard D2O sample. AND MOST
IMPORTANTLY, LOGOFF.
If you fail to do this, Admin will delete your files and revoke
your password.
ICON has been configured to send your data automatically to
the email address you will provide to Admin. This auto-send
file is a single uncompressed file with the last spectrum you
have taken. It is a pure text file in a format called JCAMP-DX.
This is a universal text format file for communicating data
between different systems or computers. In contains lots of
information on how the spectrum was taken. Fortunately, most
off-line processing programs will recognize JCAMP-DX files
and handle them. We have tested MESTREC and it correctly
recognizes this format automatically. Other packages have not
been test but should work since this is an industry-wide
standard way of exchanging nmr data.
The bad news is that the data will be sent as an email
attachment with only the standard Bruker “filename” as its
title. The subject of the email will be ICONNMR_Datamail. It will have a single attachment such as
rxt-120104.dx(190kB). Note that the exp number is not
included in the attachment name. Therefore, multiple
spectra collected by you on the same day will have the
same email name! Be sure to rename them before you
save them to your local computer or you will overwrite
data! The Bruker exp number is included in the text but it
is buried. The 25th line of the file will start out
$$ ## owner= timkovich
The 26th line contains the exp number:
$$ $$ /home/timkovich/data/rxt/nmr/rxt-120104/8/audita.txt
Where the “8” is the exp number. If you accidentally overwrite
data with multiple exp’s taken on the same day, you will have to
go back, log-in and re-send your data with manual email. This is
described on the next few slides.
The topspin window with your
spectrum.
Click File, then Send To
Email window
The fields in this
window must be
completed.
See the next slide.
Email page
All the fields on this window must be filled-in for the email
sender to function properly. The pull-down menu's at the top left
will open choices. We recommend you leave the defaults “zipcompress” and “FID+RSPEC+ISPEC”. Other combinations have
not been tested and may have bugs.
Recall that “filename” is really a directory. You cannot email a
directory. Topspin will compress all the subdirectories and files
into a single zipped file that can be emailed. Sending the fid is
better than sending the spectrum, because with offline processing
programs you can then re-process your data at will.
The zip file will contain only the current exp. You must send each
final exp even though they are all subdirectories under
“filename”.
Email, cont.
The “To” and “From” fields must contain a valid email address. As
ridiculous as it seems, they can be the same address.
The subject field is up to you. But use something you will
recognize.
The mail server field must contain, lower case, the entry
smtp.ua.edu
The software expects something in every field, so type
yourself a brief message in the final text box.
Now click
Send
Message appears in Topspin status
window that the email was sent.
/tmp is a temporary directory where the file is stored before
passing it to the smtp server. Note that the email name begins
with the “filename” of Topspin. The 6 means it was the exp
number 6 sent, along with processing parameters from proc 1.
The final extension is .bzip. This may cause problems if you are
using Windows for offline processing. On your offline computer
save the email sent by Topspin to whatever directory is
convenient for you. You will need to uncompress (unzip or
extract, all mean the same) the file before you can process your
data. The Win XP unzip software may object that it does not
recognize the extension .bzip. Not true. Try just changing the
extension from .bzip to .zip. Then extract the files.
Comments on offline Processing
Instructions are beyond the scope of this basic tutorial. The freeware package MESTREC comes with its own tutorial. Some
comments about the email you sent yourself may be helpful.
Once you have extracted the single file, the directory where you
sent the extracted files will now contain another directory, in this
example labelled “kab-111604”, the same thing we have been
calling “filename”. Open this and you will find another subdirectory
labelled “6” (remember the exp number). Open this and you will find
multiple files and still another subdirectory labelled “pdata”. One
of the multiple files will be labelled “fid”. This is what you want. If
you are using the MESTREC Open command, be sure that File
Types is set to ALL Files (or MESTREC will only show you files
it created with the extension .mrc). Select fid then Open and your
raw, unprocessed fid will appear in the MESTREC window.
Remember that descriptive title that you may or may not
have added to your data? It is in your offline file, but you
will have to work a bit to get to it. Open “pdata” then “1”
then “title”. “title” is a text file with the information you
entered.
Now let's assume you have collected all the data you need and are
ready to logout. The following slides will take you through that
process.
The first thing you need to do is reclaim your sample tube and reinsert the standard D2O sample into the spectrometer. There are
several ways to do this.
You can use the menu's you already know. Log back into ICON.
Open the Inject window, click on Insert New Sample. After the
elevator brings up your sample, remove the spinner, replace your
tube with the D2O standard, use the depth gauge, then lower the
standard just as you inserted your sample at the beginning. Now,
instead of continuing with the rest of the ICON windows, just exit
ICON and proceed to the logout instructions.
Another way is shown on the next slides.
Place the cursor over the minilock display and hit the right
button
Left click here on BSMS
panel
This new window
appears.
Note the display that
knows
there is a sample inside.
Click on the grey button
labelled Lift.
The button turns green.
The elevator turns on.
You will see the status
button “missing” light
up red, then when the
sample is at the top,the
green “up” button
lights. Proceed with
exchanging the tubes.
Then click on the green
lift button to lower the
standard D2O tube.
When the sample is
down, the “down” button
turns green. You may
exit this window by
clicking here.
Now logout.
Click on File then Exit
A confirmation window opens
Click OK
You are back in the account
home page. Click on the
RedHat icon. Another window
opens. Click on the Logout
icon. Then a confirmation
window opens, Click OK.
You will see the main logon
window. Your session ends.
Deleting Files on the Spectrometer Computer
You will quickly learn that the data files are large. Even with large
capacity, the hard disk quickly fills up. It also becomes
cumbersome to find anything if there are multiple directory entries.
Therefore good citizenship dictates that you periodically delete your
old files. The ultimate responsibility for archivng your data resides
with your offline computer and is up to you.
It is probably not a good idea to delete your current data until you
have confirmed that the email transfer worked properly. Once that
has been verified, the copy on the spectrometer in your group
account is superfluous.
The next slides demonstrate the deletion steps. It is assumed you
have logged back into your group account and started Topspin. The
deletion is done with Topspin commands, not ICON commands.
In this example we will delete all kab data taken 11-15-04.
This is a typical screen after starting Topspin.
No data is open. Your group files are currently
hidden in what is called the browser panel.
You can toggle this panel open/closed by
typing Ctrl-D on the keyboard. To expand the
contents of the browser, click on this little key
icon.
A list of the users in group belmore appears.
In this example, there is only one user. In your
account there may be many. Click on the key
next to kab.
A list of all the “filenames” created by
kab appears. We want to delete all the
data within kab-111504. Left click on
this name.
The file name becomes highlighted.
Put the cursor back on the name and
right click.
A new window opens.
Left click on Delete.
Be Sure the values
here match what
you intend to
delete!
There is no reason
to save a portion of
the data set, so
leave the “entire”
button highlighted.
Click OK
Still another window
opens. Topspin is
making you work
really hard to delete
data, to prevent
accidents. Place the
cursor on the name and
left click.
The filename becomes
highlighted. This is
another chance to be sure
you are deleting the
correct data .Hitting the
Cancel button leaves the
data intact. Let's hit
Delete.
This is absolutely your last chance
to keep the data. We are sure we
want to Delete, so click OK
Wait a minute! All this clicking and the file
name is still there! Not really. Click on the key
icon next to kab to close the list.
Now click on the icon to re-open
The 11-15-04 data is really gone.
We just had to refresh the display.
Don't forget to logout.
Windows Sizing and Positioning
Admin will initially position the Topspin and Icon windows as
displayed in this tutorial. However, it is very easy to accidently
change these settings. Windows sizing is similar to Windows and
Mac systems. Place the cursor on the window edge, click, hold and
drag to change the size.
Positioning can be tricky, because there are several modes. The one
most similar to Windows is to first place the cursor on the top blue
title bar of any open window. Right click. A submenu appears. Left
click on Move. Now the window is locked to the cursor. Move it
where you wish. A second left click frees the window and leave it in
the new location.
This is the acquisition status bar. It is useful to
have it displayed. It can be toggled on/off.
Currently on, it can be toggled off by placing
the cursor here and right clicking
This message appears. Place the cursor
within message and left click.
The bar has collapsed. To regain the bar
left clear here and toggle the bar back on.
Disclaimer and Where Do You Go From Here
This was just a very basic tutorial that just scratches the surface of
the very complex program Topspin and its baby version ICON.
The tutorial may contain errors, which over time Admin will try to
correct and update new features. However, by following the slide
presentation at your own computer, you will see most of the
screens that come up in actual operation. This is the mechanism
we are using to train the maximum number of users in the
minimum amount of time. Do not be naïve and assume that all you
need to do is a quick read of the show, and you are qualified to
operate. You will still need hands-on training by Admin.
Your ICON account and password are your Driver's License to
use the spectrometer. This presentation is your Driver's
Manual. Study it. When you feel ready, you will be
administered a written test on the contents of the manual. You
must score a B (at least 80%) to pass the written test. Then
Admin will arrange a Driver's test with hands-on training.
Once you pass that test you will be issued a password.
A password is a privilege and can be revoked.
• Bug Report
• The lock routine will sometimes make an error and not
lock properly. This happens most often when switching
between lock solvents of widely different strengths, i.e.,
CDCl3 to/from D2O. The following slides describe what
happens and how to fix it.
This step will fail.
In the lock window, you will still see
the swept lock display.
Stop the acquisition by clicking
On Stop, then Yes
This pink strip is the Command Line. On it type
lock
Then hit enter.
Select your lock
solvent then hit
OK
• You should see the solvent change from the swept signal
to the locked signal in the lock display mini-icon. Proceed
by restarting ICON.
• Bug Report 2
• When starting TOPSPIN, a text box appears that says
something like it is already running and you are not
allowed to run more than one copy. Close the text box by
clicking on the small “X” in the upper right corner. Open
another text box
Click here to start
• In the new text box, type
• shmrm(enter)
• Try restarting Topspin.
• If this still fails, logout of your group account. At the login
screen, select the option to reboot the entire computer.
Helpful
HelpfulHint:
Hint:Stop
Stopvs.
vs.Halt
Haltcommands
Commands for Long
term Acquisitions.
The Halt vs. the stop command in
ICONNMR
The halt command will stop your acquisition
and save your data. If you hit the stop button
your experiment will end, but your data will
not be saved to your directory