Dr. Sandra van MeursBruker BioSpin Ltd, UKsandra.vanmeurs@bruker.co.uk

TopSpinTopSpin

TopSpin Training Course, 22nd November 2006

OverviewOverview

• Introduction to TopSpin

• Acquiring data through IconNMR

• Processing data

• Plotting/storing data

• Interpretation – a worked example

• Questions

What is the same?What is the same?

• Before looking at the new features it is worth mentioning what has not changed in TopSpin:

- Parameters and Data format: you can still process TopSpin data with XWINNMR (not from AVII hardware)

- Most text commands are either the same or aliased to the equivalent TopSpin command (i.e. you still type efp, …)

- IconNMR has a few new features, but broadly remains the same – automation users will not need re-training, and most will never notice any difference!

- XWINPLOT is essentially the same, so if you are already using it in xwinnmr you are ready for TopSpin Plot Editor!

What is New?What is New?

• Integrated data browser

• Command line history (and concatenation)

• Configurable interface

• Full support for handling multiple datasets

• New interactive tools for Phasing, Integration, Peak Picking,…

• Interactive guides (acquisition, processing, relaxation,…)

• Acquisition status bar and tools

• Extendable via AU programs, Python programs, etc. (including access to user interface)

• …and much much more!

Acquiring DataAcquiring Data

• Sample preparation is key

• Selecting the right experiment:

- What information do you need?

- Which experiment(s) will give you this information?

IconNMRIconNMR

Manipulating your experimentManipulating your experiment

Main types of experimentMain types of experiment

Proton Carbon HMQC/

HSQC

COSY HMBC NOE

1H 13C 1H-13C correlation

1H-1H correlation

Long range 1H-13C correlation

Through space 1H-1H correlation

1 - 5 mg 5 – 25 mg

5 –10 mg 1 – 5 mg 5 - 10 mg 1 – 5 mg

1 – 5 min 10 – 30 min

10 – 20 min

5 – 10 min 10 – 30 min 2 min – 6 hours*

OH

H H

H

H

HH

H

OH

CH2

H2C

CH3 C

OH

H HH

H

H HH

CC

C

OH

H HH

H

H HH

CC

C

OH

H HH

H

H HH

CC

H

H

7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 ppm

3.15

3.00

2.08

1.03

1.00

Example of a PROTON spectrumExample of a PROTON spectrum

3Ht J = 7.1 Hz

3HddJ =1.7, 6.9 Hz

2HqJ =7.1 Hz

1HdqJ =1.7, 15.5 Hz

1HdqJ = 6.9, 15.5 Hz

Me O Me

O1 2

3

5 64

2 3 15 6

Example of CARBON spectraExample of CARBON spectra

C13DEPT45

170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 ppm

170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 ppm

170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 ppm

170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 ppm

C13CPD

C13DEPT135

C13DEPT90

C13DEPTQ

4 5

1/6

2/3 2/3

Me O Me

O1 2

3

5 64

ppm

1.01.52.02.53.03.54.04.55.05.56.06.57.07.5 ppm

1.0

1.5

2.0

2.5

3.0

3.5

4.0

4.5

5.0

5.5

6.0

6.5

7.0

7.5

Example of a COSY spectrumExample of a COSY spectrum

Me O Me

O1 2

3

5 64

2 3 15 6

ppm

1.01.52.02.53.03.54.04.55.05.56.06.57.07.5 ppm

140

120

100

80

60

40

20

0

Example of an HSQC spectrumExample of an HSQC spectrum

Me O Me

O1 2

3

5 64

2 3 15 6

Example of an HMBC experimentExample of an HMBC experiment

ppm

1.01.52.02.53.03.54.04.55.05.56.06.57.07.5 ppm

200

180

160

140

120

100

80

60

40

20

0

Me O Me

O1 2

3

5 64

2 3 15 6

An introduction to processingAn introduction to processing

• Most spectra run through IconNMR will be processed

(and perhaps printed) already

• You could simply open the processed spectrum and look at the data

• You could reprocess - fundamentally:

- multiplier, Fourier transform and phase (more on this later)

• Then you can integrate, peak pick, calibrate etc.

Data Display ButtonsData Display Buttons

Vertical position

Hz or ppm

Horizontal position

Vertical scale Horizontal scale

grid

• Buttons seen will depend upon data-type (1D or 2D), and any customisation!

Measure distance

Retain scale

Last expansion

Y-scale

All

File handling, print,copy/paste, last 1D/2D/3D

datasets

See later…

Processing 1D dataProcessing 1D data

• Processing guide (automatic/manual)

- Processing -> Data Processing Guide

• Command line automatic: “xaup” – this will probably also print a spectrum

• Command line manual: em, ft, apk/apks

• Button: if you have one set up….

…of course you could set one up…

Processing 2D dataProcessing 2D data

• Processing guide (automatic/manual)

- Processing -> Data Processing Guide

• Command line automatic: “xaup” – this will probably also print a spectrum

• Command line manual: xfb, abs1, abs2

• Button: if you have one set up….

… of course you could set one up …

Other Buttons – interactive menusOther Buttons – interactive menus

phase correction

calibration

baseline correction

peak picking

integration

multiple display

zoom overview

OutputOutput

• Printer icon

• File -> Print

• Ctrl P

“screen dump”

Worked exampleWorked example

0.60.70.80.91.01.11.21.31.41.51.61.71.81.92.02.12.22.32.42.5 ppm

6.2

3

3.1

9

2.0

3

1.0

7

1.0

3

1.0

0

1.0

0

1.0

011H spectrumH spectrum

DMSO

O

CH3

CH3CH3

H

H

H

H

H

H H12

3

4

5

6

7

A B

C

ppm

0.81.01.21.41.61.82.02.22.4 ppm

10

15

20

25

30

35

40

HSQCHSQC

CHCH3

CH2

O

CH3

CH3CH3

H

H

H

H

H

H H12

3

4

5

6

7

A B

C

HSQC expansionHSQC expansion

ppm

1.81.92.02.12.22.32.4 ppm

41.5

42.0

42.5

43.0

43.5

44.0

CHCH3

CH2

1

O

CH3

CH3CH3

H

H

H

H

H

H H12

3

4

5

6

7

A B

C

COSYCOSY

ppm

1.21.41.61.82.02.22.4 ppm

1.2

1.4

1.6

1.8

2.0

2.2

2.4

1

23 5 4

6 7

O

CH3

CH3CH3

H

H

H

H

H

H H12

3

4

5

6

7

A B

C

HMBCHMBC

ppm

0.81.01.21.41.61.82.02.22.4 ppm

10

15

20

25

30

35

40

45

50

55

12

35 4

67

CHCH3

CH2 C

C

O

CH3

CH3CH3

H

H

H

H

H

H H12

3

4

5

6

7

A B

C

2D NOESY2D NOESY

ppm

0.70.80.91.01.11.21.31.41.51.61.71.81.92.02.12.22.32.4 ppm

0.8

1.0

1.2

1.4

1.6

1.8

2.0

2.2

2.4

1

2

3 5 46 7C

2D NOESY expansion2D NOESY expansion

ppm

1.51.61.71.81.92.02.12.22.32.4 ppm

0.75

0.80

0.85

0.90

0.95

1.00

1

23 5 6

C

A

B

O

CH3

CH3CH3

H

H

H

H

H

H H12

3

4

5

6

7

A B

C

1D selective NOESY1D selective NOESY

0.70.80.91.01.11.21.31.41.51.61.71.81.92.02.12.22.3 ppm

1

23 5 46 7

C B

A

O

CH3

CH3CH3

H

H

H

H

H

H H12

3

4

5

6

7

A B

C

Interpretation summaryInterpretation summary

• Techniques

13C NMRDEPT variations1H NMRCOSYHSQC/HMQCHMBCNOESY (1D and 2D)

• Key Uses

Chemical environment, carbon countMultiplicity determinationChemical environment, quantification Spin-spin coupled nucleiDirectly bonded 1H and 13C1H and 13C correlation across multiple bonds Interactions through space

SummarySummary

• Think about data you need … don’t waste instrument time by submitting every sample for all experiments

• Remember sometimes you can obtain C13 chemical shifts from combination of 2D spectra when sample is limited

• Use processing guides !!

• Remember you can create a button for tasks you often repeat

• “Save” is implicit in TopSpin – any processing changes you make will be remembered for next time

• “Prnt” gives you quick and dirty screen dump

• Plot Editor gives you pretty spectra for hard/electronic copy

• ………….any questions??