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©2015 Waters Corporation 1
Enhancing Separations Performance for Chemicals and Polymers
Tim J Jenkins Ph.D
Director, Business Development
©2015 Waters Corporation 3
Renewed Focus on Polymer Analysis
Ion mobility enabled HDMS
Comprehensive 2D Separations
Thermal Analysis & Rheology
APC : Polymer MWD
©2015 Waters Corporation 4
Evolution of Separation Technology
Gas Chromatography Convergence Chromatography
GC
Capillary GC
HPLC
UPLC
SFC
UPC2
Liquid Chromatography
©2015 Waters Corporation 5
Definitions
Convergence Chromatography is a category of separation science
that provides orthogonal and increased separation power,
compared to liquid or gas chromatography, to solve separation
challenges.
UltraPerformance Convergence Chromatography [UPC2] is a
holistically designed chromatographic system that utilizes
liquid CO2 as a mobile phase to leverage the chromatographic
principles and selectivity of normal phase chromatography
while providing the ease-of-use of reversed-phase LC.
©2015 Waters Corporation 6
Driving UPC2 performance
–Selectivity that can be addressed
–Management of supercritical fluids
–Innovative chemistries
©2015 Waters Corporation 7
What Drives UPC2 Performance? - Addressable Selectivity
Solvent Pentane, Hexane, Heptane
Xylene
Toluene
Diethyl ether
Dichloromethane
Chloroform
Acetone
Dioxane
THF
MTBE
Ethyl acetate
DMSO
Acetonitrile
Isopropanol
Ethanol
Methanol
Stationary Phase
Silica / BEH
2-ethylpyridine
Cyano
Aminopropyl
Diol
Amide
PFP
Phenyl
C18 < C8
Reversed-phase
Selectivity Space
Normal Phase Selectivity
Space
©2015 Waters Corporation 8
Convergence Chromatography Selectivity Space
What Drives UPC2 Performance? - Addressable Selectivity
Solvent Pentane, Hexane,
Heptane
Xylene
Toluene
Diethyl ether
Dichloromethane
Chloroform
Acetone
Dioxane
THF
MTBE
Ethyl acetate
DMSO
Acetonitrile
Isopropanol
Ethanol
Methanol
Stationary Phase
Silica / BEH
2-ethylpyridine
Cyano
Aminopropyl
Diol
Amide
PFP
Phenyl
C18 < C8
Wea
k Str
ong
Supercritical CO2
Organic Modifier
©2015 Waters Corporation 9
What Drives UPC2 Performance? - Managing Supercritical Fluids
Historical challenges for SFC Lack of robustness
– Shifting retention times – Low accuracy for partial loop injections – Unstable modifier delivery at low percentages of co-solvent (< 5%)
Lack of instrument performance – Insufficient instrumentation reliability (pumping system, injection
mechanism, backpressure regulator) – Large system dispersion and dwell volumes prevented adoption of
smaller particles and high throughput analysis
Low sensitivity – High detector and pump noise – Refractive index effects of CO2
©2015 Waters Corporation 10
What Drives UPC2 Performance? - Innovative Chemistries
ACQUITY UPC2 BEH 2-EP • Good retention, peak shape and selectivity • Lipids, steroids, pesticides
ACQUITY UPC2 BEH
• Heightened interaction with polar groups such as phospholipids • OLED’s, polymer additives, pesticides
ACQUITY UPC2 CSH Fluoro-Phenyl • Good retention of weak bases • Alternate elution for acidic and neutral compounds • Vitamin D metabolites, steroids, natural products
ACQUITY UPC2 HSS C18 SB • Reversed-phase-like selectivity • Fat soluble vitamins, lipids (Free fatty acids)
Scalable to larger particle sizes ACQUITY UPC2/Viridis BEH (1.7, 3.5 and 5 µm) ACQUITY UPC2/Viridis BEH 2-EP (1.7, 3.5 and 5 µm) ACQUITY UPC2/Viridis CSH Fluoro-phenyl (1.7, 3.5 and 5 µm) ACQUITY UPC2 HSS C18 SB (1.8 and 3.5 µm)
©2015 Waters Corporation 11
Built upon proven UPLC Technology – Quantifiable increase in productivity – Ultra-low dispersion enable the use of small
diameter particles
Exceptional increase in available selectivity – Solve routine & complex separation challenges
2012 Pittcon Editors Gold Award 2013 Green Innovation Award 2013 Best New Separation Product 2013 R&D 100 Award
©2015 Waters Corporation 12
ACQUITY UPC2 Flow Path and Components
Inject valve
Auxiliary Inject valve
Column Manager
PDA detector
Back Pressure Regulator (Dynamic and Static)
Waste Modifier CO2 Supply CO2
Pump Modifier
Pump
mixer Thermo-electric heat exchanger
Make-up Pump
Mass Spec
Splitter
©2015 Waters Corporation 13
UPC2 Configuration for MS
UV Detector Make-Up Pump
Convergence Manager MS
©2015 Waters Corporation 14
The Key Benefits of UPC²
Simplify the workflow with UPC2
– Combine multiple techniques (LC & GC into CC) – Access robust normal phase separations – Eliminate solvent exchange steps for organic extracts
Deal with compound Similarity challenges – Chiral Separations (enantiomers & diastereomers) – Positional isomers (differ in location of functional groups)
Deliver Orthogonal separations
– Different relative retention helps ensure full characterization – Check method specificity by comparison to a second
procedure – Reveal “hidden” impurity or degradation peaks – Increase confidence in characterization of complex samples
SIMPLICITY
SIMILARITY
ORTHOGONALITY
©2015 Waters Corporation 16
UPC2 Polymer Separations:- Polystyrene
AU
0.00
0.20
0.40
0.60
0.80
AU
0.00
0.20
0.40
0.60
0.80
AU
0.00
0.02
0.04
0.06
0.08
0.10
0.12
Minutes0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
Column: Waters ACQUITY UPLC HSS Cyano 1.8 um, 3x100 mm. Gradient: 10 %B to 45 %B over 3 min, hold for 30 s, back to 10 % B in 30 s Flow rate: 1.7 mL/min, ABPR: 2800 psi
PS-1000
PS-2500
PS-1300
©2015 Waters Corporation 17
UPC2 Polymer Separations:- PMMA
AU
-0.002
0.000
0.002
0.004
0.006
0.008
0.010
0.012
0.014
Minutes1.00 2.00 3.00 4.00 5.00 6.00
AU
0.000
0.002
0.004
0.006
0.008
Minutes2.00 4.00 6.00 8.00 10.00 12.00 14.00
Column: Waters ACQUITY UPLC HSS Cyano 1.8 um, 3x100 mm. Flow rate: 2.0 mL/min, ABPR: 2000 psi. Gradient: 5 %B to 30 %B over 15 min, hold for 1 min, back to 5 %B in 0.1 min
PMMA-2500
PMMA-4250
Column: Waters ACQUITY UPLC HSS Cyano 1.8 um, 3x100 mm. Flow rate: 2.0 mL/min, ABPR: 2000 psi Gradient: 5 %B to 25 %B over 5 min, then to 30 %B over 1 min, back to 5 %B in 0.1 min.
©2015 Waters Corporation 18
UPC2 Polymer Separations:- Triton X-100
Non-ionic surfactant: used in cosmetics, industrial materials, and as detergents
Composition requires monitoring – Differences in ethoxy chain length affect viscosity, solubility, polarity,
and other characteristics of the product
NP-HPLC
HT-GC
SFC
©2015 Waters Corporation 19
UPC2 Polymer Separations:- Triton X-100
System: ACQUITY UPC2 with UPC2PDA Column: ACQUITY UPC2 BEH 2.1mm x 50mm, 1.7µm Mobile Phase: A: CO2 B: Methanol Wash Solvents: 70:30 Methanol:Isopropanol Separation Mode: Gradient starting at 2% B to 35% over 1.25 minutes, back to 2% B in 5 s Flow Rate: 2.0 mL/min ABPR: 1500 psi Column Temp.: 40°C Injection Volume: 1.0 µL Run Time: 2 minutes Detection: PDA 3D Channel: PDA, 210-400nm; PDA 2D Channel: 222nm @ 4.8nm Resolution (Compensated 380-480nm) CDS: Empower® 3 CDS
©2015 Waters Corporation 20
Peak1 -
0.2
71
Peak2 -
0.4
55
Peak3 -
0.5
74
Peak4 -
0.6
62
Peak5 -
0.7
33
Peak6 -
0.7
92
Peak7 -
0.8
41
Peak8 -
0.8
84
Peak9 -
0.9
21
Peak10 -
0.9
54
Peak11 -
0.9
85
Peak12 -
1.0
13
Peak13 -
1.0
41
Peak14 -
1.0
67
Peak15 -
1.0
93
Peak16 -
1.1
19
Peak17 -
1.1
58
Peak18 -
1.1
88
Peak19 -
1.2
17
Peak20 -
1.2
48
AU
0.00
0.10
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
1.10
1.20
1.30
Minutes0.10 0.15 0.20 0.25 0.30 0.35 0.40 0.45 0.50 0.55 0.60 0.65 0.70 0.75 0.80 0.85 0.90 0.95 1.00 1.05 1.10 1.15 1.20 1.25 1.30 1.35 1.40 1.45 1.50
Triton X 100
UPC2 Polymer Separations:- Triton X-100
• CO2 and Methanol gradient @ 40°C • 75s to elute all components • Approx. 20 oligomers separated and detected
1.5 mins
©2015 Waters Corporation 21
Column Name: SampleName: triton h/e 65 deg 3-20 gr 20 mni Date Acquired: 8/16/2012 8:44:13 AM EDT Instrument Method Id: 8156 Injection Id: 8174
AU
0.000
0.010
0.020
0.030
0.040
0.050
0.060
0.070
0.080
0.090
0.100
UPC2 Polymer Separations:- Triton X-100
Optimise separation for resolution (20 mins run time) See significant fine structure Ability to detect and monitor minor components and by-products
©2015 Waters Corporation 22
UPC2 Polymer Separations:- Condensation co-polymer synthesis
Bisphenol A and formaldehyde condensation co-polymer
Formaldehyde
Bisphenol A
Resin “Trimer”
©2015 Waters Corporation 23
UPC2 Polymer Separations:- Condensation co-polymer synthesis
AU
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
0.40
0.45
Minutes0.00 0.50 1.00 1.50 2.00 2.50 3.00
Unknown m/z 227
Dimer m/z 467
Trimer m/z 707
UPC2 with UV and MS detection
©2015 Waters Corporation 24
UPC2 Polymer Separations:- Condensation co-polymer synthesis
AU
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
Minutes0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50
Combined - SQ 1: MS Scan 1: 200.00-2000.00 ES-, Centroid, CV=Tune
227.0
271.0
Inten
sity
0
50000
100000
150000
200000
250000
300000
350000
400000
450000
500000
m/z200.00 250.00 300.00 350.00 400.00
Sample Mass spectrum
Bisphenol-A standard
Mass spectrum
©2015 Waters Corporation 25
UPC2 Polymer Separations:- Poly [Phenylglycidyl ether–co-formaldehyde]
Potentially complex mixture of oligomers depending on where linkage occurs between the units.
3 Potential dimers are shown below
Phenylglycidyl ether
+ Formaldehyde
Dimers
(1)
(2)
(3)
©2015 Waters Corporation 26
UPC2 Polymer Separations:- Poly [Phenylglycidyl ether–co-formaldehyde]
Dim
er 1
Trim
er 1
Dim
er 2
D
imer
3
Trim
er 2
Tr
imer
3
Trim
er 5
Tr
imer
4
Trim
er 6
Trim
er 7
UPC2 with UV detection
©2015 Waters Corporation 27
UPC2 Polymer Separations:- Poly [Phenylglycidyl ether–co-formaldehyde]
Inte
nsity
0.0
5.0x107
1.0x108
1.5x108
2.0x108
Minutes0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00 12.00 13.00 14.00 15.00 16.00 17.00
Dimers m/z 312
Trimers m/z 474 Tetramers
m/z 636
m/z 404 m/z
402
m/z 404 m/z 402
UPC2 with MS detection
©2015 Waters Corporation 29
“Hot Topic” – Extractables & Leachables
Focus from .. Packaging and Coatings Industries Consumer Product Manufacturers Cosmetics and Personal care Producers
Focus on … Extractables Leachables Intentionally added substances (IAS) Non-intentionally added substances (NIAS)
Focus because … Regulation
©2015 Waters Corporation 30
Typical extractables, IAS and NIAS
Impurities in starting materials
Chemical additives, plasticizers, antioxidants and contaminants present in individual polymers
Monomers & oligomers from incomplete polymerization
Volatile compounds from secondary packaging; inks, adhesives
Residual compounds from the surfaces of the molding equipment, antistatics etc
©2015 Waters Corporation 31
Controlled extraction study
4 material types – High Density Polyethylene bottle (HDPE) – Low Density Polyethylene container (LDPE) – Ethylene Vinyl Acetate plasma bag (EVA) – Polyvinyl Chloride blister pack (PVC)
3 different extraction solvents /
techniques – Water (Conventional oven) – Isopropanol (with Microwave and ASFE) – Hexane (with Microwave and Soxhlet)
3 separation techniques – GC – UPLC – UPC2 Apps Note 720004509en.pdf
Apps Note 720004490en.pdf
©2015 Waters Corporation 32
Irgafos 168
5-chloro-2-hydroxy-4-methylbenzophenone (5-Cl-2-OH-4-methyl BP)
Diphenyl phthalate Uvitex OB
Irganox 245
Irganox 1076
4-hydroxy-2-octyloxybenzophenone (4-OH-2-octyloxy BP)
Tinuvin 328 Irganox 1010 Lowinox 44B25
Irganox 1330
BHT
Naugard 445
Tinuvin P
Structures of Polymer Additives Investigated
©2015 Waters Corporation 33
4 min separation by UPC2 vs. 9.5 min by UPLC
UPC2
BH
T
5-Cl-
2-O
H-4
-met
hyl B
P
Tinu
vin
P
Tinu
vin
328
Irga
fos
168
2-O
H-4
-oct
ylox
y BP
Irga
nox
1076
Dip
heny
lpht
hala
te
Uvi
tex
OB
Nau
gard
445
Irga
nox
1330
Irga
nox
1010
Ir
gano
x 24
5 Lo
win
ox 4
4B25
AU
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
0.40
0.45
0.50
Minutes 0.00 0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
5-Cl-
2-O
H-4
-met
hyl B
P
UPLC
Tinu
vin
P
Dip
heny
l pht
hala
te
BH
T
Irga
nox
245
Low
inox
44B
25
4-O
H-2
-oct
ylox
y BP
Uvi
tex
OB
Nau
gard
445
Tinu
vin
328
Irga
nox
1076
Irga
nox
1330
Irga
nox
1010
Irga
fos
168 A
U
-0.15
-0.10
-0.05
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
0.40
0.45
0.50
0.55
0.60
Minutes 0.00 0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00 5.50 6.00 6.50 7.00 7.50 8.00 8.50 9.00 9.50 10.00
Chromatographic separations
©2015 Waters Corporation 34
Irgafo
s 1
68
Irganox 1
076 Ir
ganox 1
010
AU
-0.008
-0.006
-0.004
-0.002
0.000
0.002
0.004
0.006
0.008
0.010
0.012
0.014
0.016
0.018
0.020
0.022
0.024
0.026
0.028
0.030
Minutes0.00 0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00 4.20 4.40 4.60 4.80 5.00
UV chromastogram of LDPE SFE extract analysed by UPC2
©2015 Waters Corporation 35
Inte
nsity
0.0
5.0x105
1.0x106
1.5x106
2.0x106
2.5x106
3.0x106
3.5x106
Minutes0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
Combined - SQ 1: MS Scan 1: 200.00-1200.00 ES+, Centroid, CV=Tune
475.5
476.5 531.6
548.6
549.6
553.6
554.6 569.5
Inte
nsity
0.0
2.0x106
4.0x106
6.0x106
8.0x106
1.0x107
1.2x107
1.4x107
1.6x107
1.8x107
2.0x107
m/z450.00 460.00 470.00 480.00 490.00 500.00 510.00 520.00 530.00 540.00 550.00 560.00 570.00 580.00 590.00 600.00
Confirmation of identity using MS
Inte
nsity
0.0
2.0x106
4.0x106
6.0x106
8.0x106
1.0x107
Minutes0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00
Combined - SQ 1: MS Scan 1: 200.00-1200.00 ES+, Centroid, CV=Tune
475.5
476.5 531.6
548.6
549.6
553.6
554.6 572.6
Inte
nsity
0.0
2.0x106
4.0x106
6.0x106
8.0x106
1.0x107
1.2x107
1.4x107
1.6x107
1.8x107
2.0x107
2.2x107
m/z450.00 460.00 470.00 480.00 490.00 500.00 510.00 520.00 530.00 540.00 550.00 560.00 570.00 580.00 590.00 600.00
Peak in LDPE extract
Peak from Irganox 1076 std
©2015 Waters Corporation 36
Workflow Benefit of ACQUITY UPC2 for the Analysis of Polymer Extracts
Non-Polar Solvent Extraction
Polar Solvent Extraction
Inject direct on GC
Evaporate and reconstitute in a more polar solvent for
LC injection
Inject direct on LC
Back-extract with a non-polar solvent for GC injection
Polar or Non-Polar Extraction
Inject direct On UPC2
©2015 Waters Corporation 37
Analytical Scale SFE (MV-10 ASFE) Advantages of SFE
Little to No Residual Solvents Superior Yield and Purity Lower Operating Costs Processes Thermolabile Compounds Safe Scalable Variable solvent power (Tunable) Gas-like mass transfer Zero surface tension SC-CO2 has high affinity with organic solvents
©2015 Waters Corporation 38
Analytical Supercritical Fluid Extraction
Low IPA SFE
High IPA SFE
Soxhlet
Microwave
Column Name: 2-EP SampleName: LDPE high IPA SFE Date Acquired: 9/6/2012 9:28:52 PM EDT Instrument Method Id: 1953 Injection Id: 2310
AU
-0.005
0.000
0.005
0.010
0.015
Column Name: 2-EP SampleName: LDPE low IPA SFE Date Acquired: 9/6/2012 8:41:45 PM EDT Instrument Method Id: 1953 Injection Id: 2266
AU
-0.006
-0.004
-0.002
0.000
0.002
0.004
0.006
Column Name: 2-EP SampleName: LDPE IPA sox Date Acquired: 9/6/2012 6:38:07 PM EDT Instrument Method Id: 1953 Injection Id: 2151
AU
-0.005
0.000
0.005
0.010
0.015
0.020
Column Name: 2-EP SampleName: LDPE IPA mw Date Acquired: 9/6/2012 5:50:59 PM EDT Instrument Method Id: 1953 Injection Id: 2107
AU
-0.005
0.000
0.005
0.010
Minutes0.00 0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00 4.20 4.40 4.60 4.80 5.00
©2015 Waters Corporation 40
Expanding selectivity with Convergence Chromatography (CC) – Principles of CC
Understanding the technology and the instrument
Getting started with CC Method development Applications
[Click Here] for Web Overview
Order a Paper Copy [Here]
Available Now Convergence Chromatography Primer