1. Mass Spectrometry Facility
Advanced Analysis Centre
University of Guelph
Presented by:
Dr. Dyanne Brewer
Mass Spectrometry Facility Manager
May 2014

2. MS Sample Submission Workflow
Data Interpretation
Sample Submission
Sample Preparation
Data Acquisition
Training Available:
GC-MS derivitization
Protein digestion
GC-MS
LC-MSn
Metabolomics software
Proteomics software
Submission Forms:

3. Instrumentation Agilent GC-MS Bruker GC-MS-MS Bruker LC-MSn
Solid, liquid, gas, SPME
Bruker LC-MSn
Agilent LC-HR
MS-MS
GC- gas chromatography
LC- liquid chromatography
MS- mass spectrometry

4. Sample Type: Small Molecule
Volatile/semi-volatile
GC separation
MS confirmation
MS-MS quantitation
Non-volatile
LC Separation
MS confirmation/formula
MS-MS characterize/quantitate

5. Small Molecule GC-MS Sample Collection Metabolite Extraction
Compound Derivitization
Gas Chromatogram
NIST match to Glycine
N,N-bis(trimethylsilyl) trimethyl ester
Dr. Mutch UoG HHNS

6. Small Molecule Mass Accuracy
Best
ID Source
Formula
Species
m/z
Score
Diff (ppm)
Score (MFG)
TRUE
MFG
C49 H54 Cl5 N8 O9 P
(M+H)+
94.53
-0.33
Dr. Manderville UoG Chem

7. Small Molecule LC-MS Dr. Kirakosyan Un. MichiganUV
520 nm
Pelargonidin
271
Cyanidin
287
Peonidin
301
Delphinidn
303
Malvidin
331
Petunidin
317
Anthocyanins have been reported to be strong antioxidants, as well as potential anti-inflammatory and anticancer agents [3]. In addition, they act as a vasoprotectors [4] and possibly have anti-obesity effects [5,6].
tart cherry intake reduces blood lipids, plasma and tissue inflammation.
-peonidin
-hexose
-deoxihexose
Dr. Kirakosyan Un. Michigan

8. Small Molecule LC-MS Tissue Extract Dr. Kirakosyan Un. Michigan Heart
Bladder
kidney
Brain
Tissue Extract
Calibration Curve of Peonidin 3-O-glucoside
sample name
RT [min]
Area
concentration_peonidin dihexoside_fg/ul
1%heart_1.54g_(5)
10.2
42548
32.6
1%Bladder_0.75g_(1)
12946
3.2
1%kidney_1.4g_(14)
91933
81.7
1%brain_0.94g-(9)
14337
4.6
It is known that health-enhancing properties of anthocyanins cannot be fully understood without comprehensive analysis of their metabolism and tissue distribution. Our working hypothesis is that intake of tart cherry powder for about two weeks will 1) cause tissue deposition of tart cherry anthocyanins in several organs (heart, kidney, liver, brain, bladder, skeletal muscle, prostate, testes and adipose tissues) and 2) some conjugates (sugar moieties) of anthocyanins may play important roles for tissue distribution. We will therefore, confirm and identify the presence of tart cherry metabolites in targeted tissues of rats and also unravel possible post-adsorptive modifications of anthocyanins.
Dr. Kirakosyan Un. Michigan

9. Metabolomics Workflow
Separate and Detect MS
GC-MS/LC-MS
Compare
and Stats
MS database search targets
Compare to database/standard
Target MS/MS
MassHunter/MPP software
In-house Metlin/NIST library or online ID
GC-MS data
Pathway?

10. Sample Type: Large Biomolecule
Protein
Intact characterization
Clean-up LC
MS confirmation
Identification
Solution digest
In-gel digest LC
Auto MS-MS
Database search/de novo sequencing
PTM id

11. Protein MW Protein solution Clean-up LC Deconvoluted data Raw data
27, amu
Dr. Brown UoG Physics

12. Protein Identification
Protein digest
LC-Chromatogram
Protein Group
Protein ID
Accession
-10lgP
Coverage (%)
#Peptides
#Unique
PTM
Avg. Mass
Description 1
6181
Colicin_E1_Q372C
283.51 69 60 N
21734
Peptide Fragmentation
Protein Identification
Dr. Merrill UoG MCB

13. Advanced Analysis Centre
Mass Spectrometry Facility
Advanced Analysis Centre
University of Guelph
Manager: Dr. Dyanne Brewer
Research Associate: Dr. Armen Charchoglyan
Location: Science Complex Rm 1205
Phone: x58649 lab x53793 DB office