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ACE®
Excel™ UHPLC Columns
Advanced Chromatography Technologies Ltd
www.ace-hplc.com
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
Outline
Introduction
Key Product Points
Examples of Isocratic and Gradient HPLC UHPLC
Key Product Characteristics
Fixtures and Fittings
2
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
Key ACE®
Excel™ Features
Available as NEW high efficiency, ultra-inert 2mm silica particles
and suitable for UHPLC at 1000bar (15000psi)
Nine selectivities – including the unique C18-AR and C18-PFP
Highly reproducibility: column-to-column and batch-to-batch
Ultra-robust phases: NEW low dispersion column hardware and
NEW High Stability Column (HSC™) packing technology
Engineered with lower back pressures compared to other <2mm
phases due to 2mm particle size and frit technology
Fully scalable to ACE® 3mm, 5mm and 10mm
(Usable at lower pressures giving a performance boost to HPLC) Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
Waters XBridge 5 C18150 x 4.6 mm1.00 ml/min
Waters ACQUITY 1.7 BEH C1850 x 2.1 mm0.21 ml/min
HPLC: 5µm C18tr4=13.60 mins
(L/dp = 3)
UHPLC: <2µm C18tr4=4.69 mins
(L/dp = 2.9)
UHPLC: ACE Excel 2µm C18-PFPtr4=0.66 mins
Rs = 1.9
ACE Excel 2 C18-PFP
30 x 2.1 mm1.30 ml/min
Aim: obtain Rs
≥ 1.7 in shortest possible time for mixture
Sample: 1) 1,2,dimethoxybenzene, 2) 1,3-dimethoxybenzene, 3) 1,3,5-trimethoxybenzene, 4) toluene (reference).Mobile phase 50:50 MeOH / H2O; Temperature 40°C
To maintain Rs and
reduce run time,
keep L / dp ratio
constant
Using UHPLC and selectivity, it
is possible to dramatically improve
resolution allowing shorter columns
& increased flow rates
Using ACE®
Excel™ C18-PFP Selectivity & Throughput (Isocratic)
~ x23
Quicker
All trademarks are recognised
4
Rs = 1.7
Rs = 1.1
Rs = 3.4
1 2
3
4
1 2
3
4
2
3
4
1
1
2
3
4
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
ACE Excel 2 C18-PFP
30 x 2.1 mm2.5 ml/min, tG = 0.7 min
max pressure: 914 bar1 min cycle time
ACE 5 C18100 x 4.6 mm
1 ml/min, tG = 29 minmax pressure: 92 bar
40 min cycle time
ACE Excel 2 C1850 x 2.1 mm
0.6 ml/min, tG = 5 minmax pressure: 367 bar
9 min cycle time
1, aspirin; 2, phenacetin; 3, 1,3-dinitrobenzene ; 4, ethylbenzoate; 5, nimesulide; 6, ibuprofen; 7, indomethacin.
Rs = 1.8
13
2
5
4
7
6, 22.49 min
Rs = 1.7
2.5 5
6, 4.17 min1
2
54
7
Rs = 1.5
3
min1.5 2 2.5 3 3.5 40.5 1
1
2
34
5
6
7, 0.61 min
Rs = 1.8 Rs = 1.9
Rs = 1.7
Rs
≥ 1.7
HPLC: 5µm C1822.49 mins
UHPLC: <2µm C184.17 mins
UHPLC: 2µm C18-PFP0.61 mins
min5 10 15 20 250
250
0
0
Using ACE®
Excel™ C18-PFP Selectivity & Throughput (Gradient)
Aim: obtain Rs
≥ 1.7 in shortest possible time for mixture
~ x25
Quicker
5
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
Specifically engineered for lower UHPLC backpressures
ACE®
Excel™ Has Typically Lower Back Pressure For UHPLC
min0.5 1 1.5 2 2.5 3 3.5 4 4.5 5
3, 4
5
6
89
10
11
12
13
Agilent ZORBAX Eclipse 1.8 um XDB C18
Pmax: 540 bar
1 2
3
5
6
7
89
10
11
12
13 4
Waters ACQUITY 1.7um BEH C18
Pmax: 581 bar
1 2
1 2
3
4 5
6
7
8 9
10
11
12
13
ACE Excel 2 C18-AR
Pmax: 364 bar
1, 2
35
6
7
8 9 10
11
1213
4
Phenomenex Kinetex 1.7um C18
Pmax: 540 bar
50x2.1mm; conditions: A=5mM formic acid (aq); B=5mM formic acid in MeOH; tg= 3 to 100%B in 5 min; 0.6 ml/min; 40C; 254nm
1. Paracetamol 2. Hydrochlorothiazide 3. Methylphenylsulphoxide 4. Methylphenylsulphone 5. Aspirin 6. Phenacetin 7. 1,3-dinitrobenzene 8. 1,2,4-trimethoxybenzene 9. Ethylbenzoate 10. Nimesulide 11. Ibuprofen 12. Indomethacin 13. Mefenamic acid All trademarks are recognised
6
7
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
ACE®
Excel™ UHPLC Columns – Scalability & Reproducibility
ACE Excel 2 C18150 x 2.1mm0.21mL/min
ACE 3 C18150 x 3.0mm0.40mL/min
ACE 5 C18150 x 4.6mm1.00mL/min
ACE 10 C18150 x 21.2mm21.2mL/min
UHPLC
HPLC
Prep
LC2 310 6 754 8
MP: 35:65 v/v MeCN:0.1% TFA (aq); 22C; 254nm; 1. uracil; 2. 4-hydroxybenzoic acid; 3. acetylsalicyclic acid; 4. benzoic acid; 5. 2-hydroxybenzoic acid 6. ethyl paraben
3 µm
5 µm
10 µm
2 µm
1 2 34
5
6
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
NEW High Stability Column (HSC™) Packing Technology
Significantly Improves UHPLC Column Robustness
ACE®
Excel™ UHPLC Column Robustness
ACE® Excel™ 2 C18(with HSC™ Packing Technology)
Prototype 2 mm C18(without HSC™ Packing Technology)
1000 bar for ~2000 gradient runs Isocratic efficiency assessments every ~100 runs more demanding!
8
100x2.1mm; MPA 0.1% FA (aq); MPB: 0.1% FA in MeOH; 0.73mL/min; gradient: 20-90%B in 6 mins, 1 min hold; 3 min re-equilibration.
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
ACE®
Excel™ UHPLC Columns
pH and Temperature Stability:
For optimal column lifetime, as with ‘standard’ ACE columns,
the recommended pH operating range is 2-8
ACE Excel columns can be used outside of this pH range but
column lifetimes will vary with a number of factors (buffer,
temperature, solvents etc)*
ACE Excel columns can be operated at temperatures up to 100°C (but temperatures >60°C will reduce column lifetimes)
* For further information see FREE ACE booklet: ‘A Guide to HPLC and LC-MS
Buffer Selection’
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
ACE®
Excel™ C18-AR: Water Soluble Vitamins - Selectivity
1
2 3
4
5
6
7 ACE Excel 2 C18-AR Pmax: 209 bar
1 2, 3
4
5 6
7 PhenomenexKinetex 1.7 C18 Pmax: 315 bar
1
2 3
4
5 6
7 WatersACQUITY BEH C18 Pmax: 336 bar
min1 2 3 4 5
Agilent ZORBAXEclipse XDB-C18
1 2
3 5
6
7 Pmax: 299 bar
ACE Excel 2 C18
1
2 3
4 5, 6
7 Pmax: 216 bar
4
50x2.1mm; MP A: 20 mM KH2PO4, pH 2.7; MPB: 20 mM KH2PO4, pH 2.7 in MeOH/H2O (50:50 v/v); Gradient: 20 – 70 %B in 5 min; 0.31 ml/min; 40 °C; 205 nm1. Pyridoxine; 2. P-aminobenzoic acid; 3. Pantothenic acid; 4. Folic acid; 5. D-Biotin; 6. Cyanocobalamin; 7. Riboflavin
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
ACE®
Excel™ C18-PFP: Structural Isomers - Selectivity
ACE Excel 2 C18-PFP
16
4 7
5 2
3
P = 147 bar
Waters 1.7 ACQUITY BEH C18
1, 2, 3
6
4, 5
7
P = 237 bar
PhenomenexKinetex 2.6 C18
1, 2, 3
6 4, 5
7
P = 125 bar
min1 2 3 4 5 6 7 8 9
AgilentPoroshell 120 EC-C18
1, 2, 3
6 4, 5
7
P = 108 bar
50x2.1mm; MP: MeOH/H2O (50:50 v/v); 0.21 ml/min; 40 °C; 210/214 nm; 1. 1,2,3-trimethoxybenzene; 2. 1,2-dimethoxybenzene; 3. 1,2,4-trimethoxybenzene; 4. methoxybenzene; 5. 1,3-dimethoxybenzene; 6. 1,3,5-trimethoxybenzene; 7. toluene
1, 2, 3
6 4, 5
7
ACE Excel 2 C18
P = 150 bar
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
ACE®
Excel™ UHPLC Columns
A reusable ACE UHPLC column connector is available
Rated to 1720 bar (25,000 psi), these are suitable for at least 10
make / break cycles when correctly fitted
Suitable for use at temperature up to 100C
A torque wrench is recommended to ensure the correct force is
applied
EXL-CC = 1 / pk
EXL-CC10 = 10 / pk
EXL-TW =1 / pk
Australia & New Zealand contact: [email protected] or call +61 7 3205 5233
ACE®
HPLC / UHPLC Columns
ACE®
Excel™ UHPLC Columns
A starter kit containing 1 x Torque Wrench and 4 x reusable
connectors is available
EXL-CCSK+ +++