1h qnmr of epa and dha omega-3 fatty acid: pls-regression models obtained at 60 and 300 mhz

1H qNMR of EPA and DHA Omega-3 Fatty Acids - PLS Regression Models Obtained at 60 and 300 MHz : At-Line and On-Line

Monitroing of a Fish Oil Nutritional Supplement Manufacturing Process

John C. Edwards and Paul J. GiammatteoProcess NMR Associates, LLC, 87A Sand Pit Rd, Danbury, CT 06810 USA

Presented at the FoodMR 2014May 23, 2014Cesena, Italia

www.process-nmr.com www.nmrblog.com Tel: 203-744-5905

http://www.process-nmr.com/

http://www.nmrblog.com/

60 MHz NMR3-10 mm SamplesLab or On-Line

aspectimaging60 MHz NMR3-10 mm SamplesLab or On-Line

60 MHz NMR3-5 mm SamplesLab Only

42 MHz NMR3-5 mm SamplesLab Only

60 MHz NMR3-5 mm SamplesLab Only 60 MHz NMR

3-5 mm SamplesLab Only

45 MHz and 82 MHz NMR300 micron capillary injectionLab Only

85+ MHz NMR3-5 mm SamplesLab Only

Picospin 45 and 80

NMReady 60

Pulsar

Fourier 60MobiLab™ SPECTRA 2T

Cryogen FreePermanent MagnetHigh ResolutionBenchtop NMR

60 and 90 MHz5 mm SamplesLab Only

2 x Varian Mercury 300 Liquids1 x Varian UnityPlus-200 SolidsMultiple 10/20/40 mm TD-NMR 20-25 MHzNMR MOUSE2 x Aspect-AI 60 MHz HiRes-NMR3 x Qualion 60 MHz HiRes NMR

MicroESRShimazu GCMNova(Verify, qNMR, DB), ACDLabs

Chemometrics: GramsIQ, Eigenvector Solo, Pirouette

On-Line Process NMR

45 Systems Installed 1996-2004

Predominantly in Petroleum and Petrochemical

60 MHz Reaction Monitoring

50 sample initial data set from all points in manufacturing process.

First round of PLS regression analysis revealed concentration outliers that were linked to limitations in the GC method. This study was performed on improved GC data values.

Final 50 Sample models were utilized to validate the calibrations on a 24 sample validation set

b

b

a

g


p



Experimental Varian Mercury - 300 MHz NMR Supercon Spectrometer4 pulse on pure sample in 5 mm tube, Run UnlockedIn MNova 8.1.2 SPC Files Imported, Stacked, Binned at 3 Hz interval, Area Normalized to 100, Saved as Transposed Ascii MatrixPLS Regression Performed Thermo Grams IQ and Eigenvector Solo

300 MHz 1H NMR

DHA

EPA-a

Olefins

=C-CH2-C=

In chain

CH2

EtOOC-CH2-R

DHA

EPA

CH3-CH2-O-OC-CH2-R

CH3

EthylCH3

R-CH2-C=CH3

w6/ w9

w3EPA-b

Comparison of High EPA and High DHA PUFA-Ester after Separation

DHA

EPA

EPA

EPA

EPA

DHA

300 MHz 1H NMR Spectra of Samples at Diferent Points in the Process

-200 -150 -100 -50 0 50 100 150 200-20

-15

-10

-5

0

5

10

15

Scores on LV 1 (98.04%)

Score

s o

n L

V 2

(0.6

4%

)

Samples/Scores Plot of SPC Files in MNova.xlsx

First Esterification

Clathration

Molecular Distillate

Pollock Oil

Separator Product

PUFA Ester

Clathration Raffinate

MUFA Ester

Salmon Oil

95% Confidence Level




300 MHz 1H NMR Data

Stacked and integrated.

Integrals Utilized in PLS Regression Against EPA/DHA content by GC

FTIR-ATR Spectra

NMR Spectra

Fusion of 1H NMR and FTIR-ATR Spectra – absolute intensity scaling




PLS Regression DataEPA Content (Wt%)

R2, SECV (Wt%)DHA Content (Wt%)

R2, SECV (Wt%)

60 MHz NMR 0.988, 2.13 0.991, 1.13

300 MHz NMR 0.993, 1.68 0.992, 1.09

Peak Integral Data 0.981, 2.71 0.964, 2.24

Fused - NMR-FTIR 0.993, 1.62 0.992, 1.08

Regression Results for Various NMR Approaches




Further Steps:

Models have been expanded from 74 samples to 250 samples better covering extreme EPA and DHA content samples

Validated against 50 samples provided by customer.

Validation was passed and 50 Validation Samples were included into the models.

NMR project found problems with the EPA GC analysis that were not known to thecustomers’ laboratory and engineering personnel.

Near-Future Steps:

Laboratory system being purchased to provide rapid (2 minute analysis). Reduces GC to validation method and increases throughput of laboratory 20 fold.

On-line system will be purchased and placed into the process with a 6 stream sample system for real-time montioring.

Models will be expanded to include full fatty acid distribution of the sample by 1H NMR correlation to the full GC data provided for 300 samples.

New project on Manufacturing process of Polyalphaolefins is in final stages of feasibility and validation – expect on-line 60 MHz system in combination with TD-NMR for viscosity measurement.

Acknowledgements

Daniel EdwardsAdam Dicaprio

Generic Customer

Aspect AI – NMR SystemsCosa-Xentaur

Practical Applications of NMR in Industry ConferenceFebruary 9-12, 2015

San Diego, CAhttp://www.panicnmr.com/

http://www.panicnmr.com/

1h qnmr of epa and dha omega-3 fatty acid: pls-regression models obtained at 60 and 300 mhz

Health & Medicine

h nmr datastacked

sampleslab onlypicospin

sampleslab only45

sampleslab only42

sampleslab only60

h nmr spectra of samples

h nmr dhaepaaolefins

hiresnmr3 x qualion