Blog on Hiatus from September 21 to October 2

Due to some business travel, the blog P2C2E will be on hiatus for 2 weeks from September 21 to October 2. Posts will resume the following week.

Stay tuned for Part 2 of the series Stereochemistry Information from NOESY/ROESY data.

Blog on Hiatus from September 21 to October 2

Due to some business travel, the blog P2C2E will be on hiatus for 2 weeks from September 21 to October 2. Posts will resume the following week.

Stay tuned for Part 2 of the series Stereochemistry Information from NOESY/ROESY data.

Stereochemistry Information from NOESY/ROESY data … Part 1

Several NMR experiments offer tools to help determine the stereochemistry of a structure. Some typical experiments are 1D NOE (Nuclear Overhauser Effect), 2D NOESY (NOE Spectroscopy) and ROESY (Rotating-frame Overhauser Effect Spectroscopy). These experiments will produce signals for nuclei that are close to each other through space independent of the number of bonds separating the nuclei.

A simplified 1H-1H NOESY spectrum is shown below. The spectrum shows 2 correlations at (4.29,1.28) and (4.29,3.13) ppm. There is no correlation to the proton signal at 2.68 ppm.

Based on NOESY data, there are 2 possible conformations. The enantiomers (partially drawn) are shown below.

Stereochemistry Information from NOESY/ROESY data … Part 1

Several NMR experiments offer tools to help determine the stereochemistry of a structure. Some typical experiments are 1D NOE (Nuclear Overhauser Effect), 2D NOESY (NOE Spectroscopy) and ROESY (Rotating-frame Overhauser Effect Spectroscopy). These experiments will produce signals for nuclei that are close to each other through space independent of the number of bonds separating the nuclei.

A simplified 1H-1H NOESY spectrum is shown below. The spectrum shows 2 correlations at (4.29,1.28) and (4.29,3.13) ppm. There is no correlation to the proton signal at 2.68 ppm.

Based on NOESY data, there are 2 possible conformations. The enantiomers (partially drawn) are shown below.

Determining the Site of Modification … Part 3

Tandem mass spectrometry involves the process of selecting and separating a product ion(s) (or daughter ion(s)) and fragmenting it in a second mass analyzer. This is commonly referred to as MS/MS or MS2. Additional tandem processes can be applied to ions in the MS/MS data to create MS3 data, and so forth.

The metabolites A and B share the same exact mass, and as such, cannot be differentiated by the MS data alone (see Part 1 of this series). The MS/MS data, described in Part 2, offers fragment information that can assist in eliminating one of the candidates. Taking it a step further and thus verifying the candidate metabolite A, ESI+ MS3 data is presented herein.

The fragmentation scheme below shows both the Parent and Metabolite A with the fragment at 121 Da, in bold, fragmenting to create a fragment at 93 Da. This is supported by the nearly-identical MS3 data displayed below.

 

On the right-hand side of the fragmentation scheme, the fragments 106 and 108 Da are expected for the MS3 data of the Parent and Metabolite A, respectively.

Determining the Site of Modification … Part 3

Tandem mass spectrometry involves the process of selecting and separating a product ion(s) (or daughter ion(s)) and fragmenting it in a second mass analyzer. This is commonly referred to as MS/MS or MS2. Additional tandem processes can be applied to ions in the MS/MS data to create MS3 data, and so forth.

The metabolites A and B share the same exact mass, and as such, cannot be differentiated by the MS data alone (see Part 1 of this series). The MS/MS data, described in Part 2, offers fragment information that can assist in eliminating one of the candidates. Taking it a step further and thus verifying the candidate metabolite A, ESI+ MS3 data is presented herein.

The fragmentation scheme below shows both the Parent and Metabolite A with the fragment at 121 Da, in bold, fragmenting to create a fragment at 93 Da. This is supported by the nearly-identical MS3 data displayed below.

 

On the right-hand side of the fragmentation scheme, the fragments 106 and 108 Da are expected for the MS3 data of the Parent and Metabolite A, respectively.

Determining the Site of Modification … Part 2

Peak matching involves the process of comparing spectral data from a parent or starting material to an unknown compound. (The unknown compound can be referred more specifically as the product, impurity, degradant, metabolite, etc.). The similarities between the data indicate regions that have not changed while the differences indicate regions of change.

The full scan MS data in Part 1 does not offer enough spectral information to eliminate one of the candidate structures. The next step is to examine MS/MS data.

The ESI+ product ion spectra (MS/MS) for both the Parent (m/z 226) and Metabolite (m/z 228) are shown below. The spectra share a common fragment at m/z 121 and differ in the fragments at m/z 134/136 and 149/151.

Based on the fragments shown below, the suspected site of hydrogenation is on the carbonyl for metabolite A.

 

Determining the Site of Modification … Part 2

Peak matching involves the process of comparing spectral data from a parent or starting material to an unknown compound. (The unknown compound can be referred more specifically as the product, impurity, degradant, metabolite, etc.). The similarities between the data indicate regions that have not changed while the differences indicate regions of change.

The full scan MS data in Part 1 does not offer enough spectral information to eliminate one of the candidate structures. The next step is to examine MS/MS data.

The ESI+ product ion spectra (MS/MS) for both the Parent (m/z 226) and Metabolite (m/z 228) are shown below. The spectra share a common fragment at m/z 121 and differ in the fragments at m/z 134/136 and 149/151.

Based on the fragments shown below, the suspected site of hydrogenation is on the carbonyl for metabolite A.