Thermo Scientific

Accucore HPLC Columns

Technical Guide

Ultimate Core Performance

to Maximize Your Investment
Core Enhanced Technology
To meet modern requirements for outstanding performance free from constraints
on the type of instruments that can be used we have developed Accucore™ HPLC
columns using an exciting new concept.

The key components of Core Enhanced Technology

Solid Core Particles Tight Control of Particle Diameter

2.6µm diameter particles with a solid core Enhanced selection process keeps particle size
generate high speed, high resolution separations distribution to a minimum and produces high
without excessive backpressure efficiency columns

Automated Packing Process Advanced Bonding Technology

Enhanced automated procedures ensure that all Optimized phase bonding creates a series of high
columns are packed with the highest quality coverage, robust phases

Solid Core Tight Control of Advanced Bonding Automated Packing

Particles Particle Diameter Technology Process
Fast Separations • • • •
High Peak Capacity • • • •
Increased Sensitivity • • • •
Lower Pressure • • •
Loading Capacity • • •
Reproducible Chromatography • • •
Long Lifetime • • • •
Wide Selectivity •

The table above shows how the components of Core

Enhanced Technology contribute to the outstanding
features of Accucore HPLC columns.
Table of Contents

Particle Evolution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2
Theory of Fast HPLC . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
Why Core Enhanced Technology Works . . . . . . . . . . . . . . . . 4
Core Enhanced Technology Effect . . . . . . . . . . . . . . . . . . . 5
Faster than 5µm and 3µm . . . . . . . . . . . . . . . . . . . . . . . . . 6
Short Columns for Even Faster Separations . . . . . . . . . . . . . . 7
Higher Peak Capacity than 5µm and 3µm . . . . . . . . . . . . . . . 8
More Sensitive than 5µm and 3µm . . . . . . . . . . . . . . . . . . . 9
Equivalent Performance to Sub-2µm with Lower Pressure . . . . 10
Loading Capacity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Simple Method Transfer . . . . . . . . . . . . . . . . . . . . . . . . 12
UHPLC System Not Required . . . . . . . . . . . . . . . . . . . . . . 13
Instrument Optimization . . . . . . . . . . . . . . . . . . . . . . . . . 14
Reproducible Chromatography . . . . . . . . . . . . . . . . . . . . 15
Long Lasting Columns . . . . . . . . . . . . . . . . . . . . . . . . . . 16
Column Protection . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18
Thermo Scientific Accucore Optimum Selectivity . . . . . . . . . . 19
RP-MS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
C18 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
aQ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22
Phenyl-Hexyl . . . . . . . . . . . . . . . . . . . . . . . . . . . 23
PFP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
HILIC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25
Applications
Food . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
Environmental . . . . . . . . . . . . . . . . . . . . . . . . . . . 27
Clinical . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28
Forensic Toxicology . . . . . . . . . . . . . . . . . . . . . . . . 30
Forensics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
Pharmaceuticals . . . . . . . . . . . . . . . . . . . . . . . . . . 31
Bio-pharmaceuticals . . . . . . . . . . . . . . . . . . . . . . . 33
General . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34
Thermo Scientific Options for Fast HLPC . . . . . . . . . . . . . . . 35
Ordering Information
Columns and Guards . . . . . . . . . . . . . . . . . . . . . . . 36
Kits . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37

Thermo Scientific Accucore HPLC Columns | Technical Guide 1

 Particle Evolution
In the search for ever faster and better separations the size and shape of column
packing materials has evolved in the decades since the invention of HPLC.
Packing materials have changed from large pellicular particles via smaller totally
porous particles to spherical particles with diameters of less than 2µm.
Our Core Enhanced Technology has changed things again. These particles
are not totally porous, but rather have a solid core and a porous outer layer.

Large, irregular particles 5µm particles 3µm particles Sub-2µm particles

Core Enhanced
Technology
particles

Solid Core

Core Enhanced Technology

Core enhanced technology offers the advantages associated with sub-2µm packing
materials – fast, high-resolution separations – without the high backpressures required
to work with the smaller particles.

2 Thermo Scientific Accucore HPLC Columns | Technical Guide

Theory of Fast HPLC
The Van Deemter curves and kinetic plot below show the impact of packing
material particle diameter and type on chromatographic efficiency.
The highest efficiencies and lowest rate of efficiency loss with increasing flow rate
are seen with the smallest particle diameter and the Core Enhanced Technology.

Fully Porous 5µm

Fully Porous 5µm
Fully Porous 3µm
Fully Porous 3µm
Fully Porous 1.9µm
Fully Porous 1.9µm
Core Enhanced
Core Enhanced
Technology 2.6µm
HH

Technology 2.6µm

On the Van Deemter curve it can be seen that plate

height (H) reaches a minimum (maximum efficiency)
at an optimum mobile phase velocity and then
µ (mm/s) decreases as velocity increases.
µ (mm/s)

Totally Porous 5µm

Totally Porous 5µm
Totally Porous 3µm
Totally Porous 3µm
Totally Porous 1.9µm
Totally Porous 1.9µm
Core Enhanced
Core Enhanced
Technology 2.6µm
Technology 2.6µm
(s)(s)
t0/N
t0/N

In this kinetic plot efficiency (N) decreases as function

Nopt
Nopt of decreasing plate generation time (t0/N) – where t0 is
the retention time of an unretained peak and inversely
proportional to flow rate. The optimal performance
(Nopt) is indicated.
N
N

Resolution Poor Resolution Good Resolution

The general chromatographic resolution equation
shows that resolution is directly proportional to the
square root of efficiency. This means that with small
diameter and core enhanced technology relatively
high flow rates can be used, whilst maintaining the
high efficiencies required for good separations.

a Selectivity
1 (a - 1) k
N Efficiency RS = √N
k Capacity factor 4 a 1+k

Thermo Scientific Accucore HPLC Columns | Technical Guide 3

 Why Core Enhanced Technology Works
Two factors that strongly affect chromatographic efficiency are mass transfer and eddy diffusion,
the C and A terms respectively from the Van Deemter equation.

B
H=A+ + Cu
u

H Height equivalent to theoretical

plate (column length/efficiency)
A Eddy diffusion
B Longitudinal diffusion
C Resistance to Mass Transfer Resistance to mass transfer is minimized by the solid core design of Core
Enhanced Technology particles as the diffusional path of analytes is limited
u Mobile phase linear velocity by the depth of the outer porous layer.

Material Accucore 2.6μm

Average Particle Size
1.12
Distribution (D90/10)

The tight control of Core Enhanced Technology particle diameter and

automated packing process used for Accucore HPLC columns result in a tight,
highly uniform packed bed that minimizes eddy diffusion.

Lower Backpressure
L Column length (cm)
h Mobile phase viscosity (cP)
250LhF
F Flow rate (mL/min) DP ~
dp2 Particle diameter (μm)
dp2dc2
dc2 Column diameter (cm)

The equation above shows how backpressure is related to particle diameter.

The tightly controlled 2.6μm diameter of Accucore particles also results in lower backpressures
than typically seen with sub-2μm materials.

4 Thermo Scientific Accucore HPLC Columns | Technical Guide

Core Enhanced Technology Effect
The plots below show how the efficiency and backpressure of Accucore HPLC columns compare
to columns packed with traditional totally porous 5μm, 3μm and 1.9μm particles.
Decreasing Efficiency

18.0 Hypersil GOLD 5µm

16.0
Hypersil GOLD 3µm
14.0
18.0 Hypersil GOLD 5µm
Hypersil GOLD 1.9µm
12.0
16.0
Hypersil GOLD 3µm
10.0
14.0 Accucore RP-MS 2.6µm
H

Hypersil GOLD 1.9µm

8.0
12.0

6.0
10.0 Accucore RP-MS 2.6µm
H

4.0
8.0

2.0
6.0

0.0
4.0
0.0 2.0 4.0 6.0 8.0 10.0 Accucore HPLC columns are more
2.0 µ (mm/s) efficient than 5μm and 3μm columns.
0.0
0.0 2.0 4.0 6.0 8.0 10.0
µ (mm/s)

Hypersil GOLD 5µm

1000
900 Hypersil GOLD 3µm
Increasing Pressure

800 Hypersil GOLD 5µm

1000 Hypersil GOLD 1.9µm
(bar)

700
900 Hypersil GOLD 3µm
600 Accucore RP-MS 2.6µm
Pressure Pressure

800
500 Hypersil GOLD 1.9µm
Typical HPLC Pressure Limit
(bar)

700
400
600
300
Accucore RP-MS 2.6µm
500
200 Typical HPLC Pressure Limit
400
100
300
0
200 0 200 400 600 800 1000
100 Flow Rate (µL/min)
0
0 200 400 600 800 1000 Accucore HPLC columns achieve this
Flow Rate (µL/min) efficiency without excessive backpressure

Thermo Scientific Accucore HPLC Columns | Technical Guide 5

 Faster than 5μm and 3μm
Using Accucore HPLC columns excellent separations can be achieved in shorter times.
The examples on this page show how by increasing flow rates while maintaining efficiency,
and therefore resolution, the time taken to separate a mixture can be reduced by a factor
of 3 and solvent costs can be reduced by 7-times!

Rs=2.64 Hypersil GOLD 5µm Mobile phase: A – Water; B – Acetonitrile

150 x 4.6mm
Gradient: Accucore RP-MS 2.6µm
100 x 2.1mm = 35 – 60% B in 3.5 minutes
Rs=1.64
Hypersil GOLD 3µm
Hypersil GOLD 5µm
100 x 2.1mm
100 x 2.1mm = 35 – 60% B in 4.0 minutes
Hypersil GOLD 5µm
mAU

Rs=1.96
100 x 2.1mm = 35 – 60% B in 6.7 minutes
Hypersil GOLD 3µm Hypersil GOLD 5µm
100 x 2.1mm 150 x 4.6mm = 35 – 60% B in 10.0 minutes
Flow rate: Accucore RP-MS
2.6µm 100 x 2.1mm = 400µL/min
Rs=2.50 Accucore RP-MS 2.6µm Hypersil GOLD 3µm
100 x 2.1mm
100 x 2.1mm = 350µL/min
0 1 2 3 4 5 6 7 8 9 10
Hypersil GOLD 5µm
Minutes 100 x 2.1mm = 210µ/min
Hypersil GOLD 5µm
150 x 4.6mm = 1000µL/min
Injection volume: 1µL (Hypersil GOLD 5µm 150 x 4.6mm = 5µL)
Temperature: 30°C
Detection: UV at 247nm (0.1s rise time, 20Hz)
Analytes: 1. Tebuthiuron
2. Metoxuron
3. Monuron
4. Chlorotoluron
5. Diuron
6. Linuron

Accucore RP-MS Hypersil GOLD Hypersil GOLD Hypersil GOLD

2.6µm, 100 x 2.1mm 3µm, 100 x 2.1mm 5µm, 100 x 2.1mm 5µm, 150 x 4.6mm
Resolution (critical pair) 2.50 1.96 1.64 2.64
Run time (min)
6.00 7.00 11.50 17.00
including gradient re-equilibration

Reducing analysis time and solvent costs results in higher

throughput and lower cost per analysis.

6 Thermo Scientific Accucore HPLC Columns | Technical Guide

Short Columns for Even Faster Separations
The separating power of Accucore HPLC columns means that by using shorter column dimensions
acceptable resolution can be maintained, with even greater increases in throughput
and reduction in costs.

Rs=2.50 Mobile phase: A – Water; B – Acetonitrile

Gradient: Accucore RP-MS 2.6µm
50 x 2.1mm = 35 – 60% B in 1.8 minutes
Accucore RP-MS 2.6µm
100 x 2.1mm = 35 – 60% B in 3.5 minutes
mAU

Accucore RP-MS 2.6µm

100 x 2.1mm Flow rate: 400µL/min

Rs=1.51

Accucore RP-MS 2.6µm

50 x 2.1mm
-100
0 0.5 1 1.5 2 2.5 3 3.5 4 4.5 5
Minutes

Analysis Time and Solvent Savings

Accucore RP-MS Accucore RP-MS A 50mm column gives

2.6µm, 50 x 2.1mm 2.6µm, 100 x 2.1mm acceptable separation
Resolution (critical pair) 1.51 2.50 with a doubling of
Run time (min)
3.00 6.00 productivity and halving
including gradient re-equilibration of solvent costs.

Thermo Scientific Accucore HPLC Columns | Technical Guide 7

 Higher Peak Capacity than 5μm or 3μm
As an alternative to speeding up analysis the high resolution offered by Accucore HPLC columns
can also be used to improve complex separations through an increase in peak capacity.

nc Peak capacity
tg
tg Gradient time nc = 1 + (W)
W Peak width (10% height)

Mobile phase: A – Water; B – Acetonitrile

Hypersil GOLD 5µm
Gradient: 65 – 95% B in 2.1 minutes,
100 x 2.1 mm
95% B for 0.4 minute
Flow rate: 400µL/min
Injection volume: 1µL
mAU

Hypersil GOLD 3µm Temperature: 40°C

100 x 2.1mm
Detection: UV at 247nm (0.1s rise time, 20Hz)
Analytes: 1. Acetophenone
2. Propiophenone
Accucore RP-MS 2.6µm 3. Butyrophenone
100 x 2.1mm 4. Valerophenone
5. Hexanophenone
0 0.5 1 1.5 2 2.5 3 6. Heptanophenone
Minutes 7. Octanophenone

Peak Capacity Comparison

170
Normalized Peak Capacity

160

150

140

130

120 Accucore RP-MS 2.6µm 158

110 Hypersil GOLD 3µm 132
100
Hypersil GOLD 5µm 100
90
Accucore RP-MS 2.6µm Hypersil GOLD 3µm Hypersil GOLD 5µm

The higher the peak capacity the more

analytes can be identified within a run.

8 Thermo Scientific Accucore HPLC Columns | Technical Guide

More Sensitive than 5μm or 3μm
According to the formula shown below, the sharper, taller peaks obtained with Accucore
HPLC columns result in a higher signal to noise ratio (S/N) and therefore better sensitivity.

cmax Concentration at peak apex

N Efficiency
√N Vi
Vi Injection volume cmax∝
L dc2 (1 + k)
L Column length
dc Column internal diameter
k Capacity factor

169 Mobile phase: A – Water; B – Acetonitrile

Hypersil GOLD 5µm
Gradient: Accucore RP-MS 2.6µm
100 x 2.1mm
100 x 2.1mm = 35 – 60% B in 3.5 minutes
Hypersil GOLD 3µm
368 100 x 2.1mm = 35 – 60% B in 4.0 minutes
Hypersil GOLD 5µm
mAU

Hypersil GOLD 3µm 100 x 2.1mm = 35 – 60% B in 6.7 minutes

100 x 2.1mm
Flow rate: Accucore RP-MS 2.6µm
399 100 x 2.1mm = 400µL/min
Hypersil GOLD 3µm
100 x 2.1mm = 350µL/min
Accucore RP-MS 2.6µm
100 x 2.1mm
Hypersil GOLD 5µm
100 x 2.1mm = 210µL/min
0 1 2 3 4 5 6 7 8 Injection volume: 1µL
Minutes Temperature: 30°C
Detection: UV at 247nm (0.1s rise time, 20Hz)
Analytes: 1. Tebuthiuron
2. Metoxuron
3. Monuron
4. Chlorotoluron
5. Diuron
6. Linuron

Sensitivity

Column S/N (6-sigma) for Monuron Increase in Sensitivity

Accucore 2.6µm, 100 x 2.1mm 399 136%
Hypersil GOLD 3µm, 100 x 2.1mm 368 117%
Hypersil GOLD 5µm, 100 x 2.1mm 169 –

Better sensitivity allows reliable detection

and determination of small peaks, for example
low level impurities.

Thermo Scientific Accucore HPLC Columns | Technical Guide 9

 Equivalent Performance to Sub-2μm
with Lower Pressure
With solid core design, tight particle size distribution and uniform packed bed
Accucore HPLC columns have broadly equivalent performance to sub-2μm columns
and yet generate only a fraction of the backpressure.

Mobile phase: A – Water; B – Acetonitrile

Gradient: 65 – 95% B in 1.7 minutes
95% B for 0.3 minute
Hypersil GOLD 1.9µm
100 x 2.1mm Flow rate: 500µL/min
mAU

Injection volume: 1µL

Temperature: 40°C
Detection: UV at 247nm (0.1s rise time, 20Hz)
Analytes: 1. Acetophenone
Accucore RP-MS 2.6µm 2. Propiophenone
100 x 2.1mm
3. Butyrophenone
4. Valerophenone
0 0.5 1.0 1.5 2.0 2.5 5. Hexanophenone
Minutes 6. Heptanophenone
7. Octanophenone

Pressure

Accucore RP-MS Hypersil GOLD

2.6µm, 100 x 2.1mm 1.9µm, 100 x 2.1mm
Resolution (critical pair) 3.72 4.20
Run time (min) 3.50 3.50
Maximum pressure (bar) 171 338

Lower backpressure eliminates the requirement for UHPLC

systems with maximum pressure ratings >600 bar. If a UHPLC
system is used then the lower backpressure reduces wear
on the instrument.

10 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Loading Capacity
With tightly packed beds and high bonded phase coverage Accucore HPLC columns have
loading capacities that allow a wide range of analyte concentrations to be determined.
The example below shows minimal change in retention and peak shape with increasing
analyte concentration.

2500000 1.2 Column: Accucore RP-MS 100 x 2.1mm

Mobile phase: 68:32 (v/v) Water/Methanol
1 Flow rate: 1.0mL/min
2000000
R² = 0.9998 Temperature: 40° C

Normalised Value
0.8 Detection: 254nm
Peak Area

1500000 Injection volume: 1µL

0.6

1000000 Asymmetry
Concentration (ng/μL) Load on Column (μg)
0.4
5 0.005 Efficiency
500000
0.2 25 0.025
50 0.050 Retention Time

0 0 250 0.250
0 0.5 1.0 1.5 2.0 2.5 0 500 1.0 0.500 2.0 3.0
Load on column (µg) Load on column (µg)
1000 1.000
2000 2.000
1.2

1
Normalised Value

0.8

0.6
Asymmetry
0.4
Efficiency

0.2
Retention Time

0
2.0 2.5 0 1.0 2.0 3.0
Load on column (µg)

Thermo Scientific Accucore HPLC Columns | Technical Guide 11

 Simple Method Transfer
Fast HPLC is often performed using lower volume columns.

A few simple steps are required to transfer a method to a lower volume Accucore HPLC column.

Method Transfer Tool

A convenient method transfer tool is available at the Chromatography Resource Center
www.thermoscientific.com/crc
• Adjust Flow Rate
Keep linear velocity constant between original and new method, taking into account
particle size and geometry
• Adjust Injection Volume
Keep the ratio of injection volume to column volume constant
• Adjust Gradient Profile
Keep the number of column volumes constant for each gradient segment

Mobile phase: A – Water; B – Acetonitrile

Gradient: Accucore RP-MS 2.6µm
Hypersil GOLD 5μm
100 x 2.1mm = 35 – 60% B in 3.5 minutes
150 x 4.6mm
Hypersil GOLD 5µm
150 x 4.6mm = 35 – 60% B in 10.0 minutes
mAU

Flow rate: Accucore RP-MS 2.6µm

100 x 2.1mm = 400µL/min
Hypersil GOLD 5µm
150 x 4.6mm = 1000µL/min
Injection volume: Accucore RP-MS 2.6µm
Accucore RP-MS 2.6µm 100 x 2.1mm = 1µL
100 x 2.1mm Hypersil GOLD 5µm
150 x 4.6mm = 5µL
0 1 2 3 4 5 6 7 8 9 10 Temperature: 30°C
Minutes
Detection: UV at 247nm (0.1s rise time, 20Hz)
Analytes: 1. Tebuthiuron
2. Metoxuron
3. Monuron
4. Chlorotoluron
5. Diuron
6. Linuron

12 Thermo Scientific Accucore HPLC Columns | Technical Guide

UHPLC System Not Required
The low backpressures generated associated with Core Enhanced Technology mean
that Accucore HPLC columns can be used with both UHPLC and HPLC systems.

Column: Accucore RP-MS 2.6µm, 100 x 2.1mm

Mobile phase: A – Water; B – Acetonitrile
Gradient: 65 – 95% B in 2.1 minutes
95% B for 0.4 minute
Thermo Scientific Accela 1250
Flow rate: 400µL/min
mAU

Injection volume: 1µL

Temperature: 40°C
Detection: UV at 247nm (0.1s rise time, 20Hz)
Analytes: Phenones
1. Acetophenone
Thermo Scientific Surveyor
2. Propiophenone
0 0.4 0.8 1.2 1.6 2 2.4 2.8 3.2 3.6 4 3. Butyrophenone
Minutes 4. Valerophenone
5. Hexanophenone
6. Heptanophenone
7. Octanophenone
mAU

Agilent 1100

0 0.4 0.8 1.2 1.6 2 2.4 2.8 3.2 3.6 4

Minutes

System Comparison
The UHPLC system gives the best performance and any HPLC system can benefit from the faster,
high resolution separations offered by Accucore HPLC columns. The higher resolution observed
with the Surveyor is caused by the gradient delay.

Accela 1250 Surveyor Agilent 1100

Run Time (min) 2.5 3.0 3.5
Average Peak Width at 50% height (min) 0.02 0.02 0.04
Average Resolution (USP) 6.15 6.53 5.33

In order to get the best out of Accucore

HPLC columns the system should be
optimized for high efficiency separations.
See Instrument Optimization on page 14.

Thermo Scientific Accucore HPLC Columns | Technical Guide 13

 Instrument Optimization Page 14

Accucore HPLC columns produce very narrow peaks. In order to preserve this efficiency
the HPLC system should be optimized to reduce any potential causes of peak broadening.
Instrument Optimization

Potential causesAccucore
of peakcolumns
broadening are:narrow peaks. In order to preserve this efficiency the HPLC system
produce very
Extra-column band should be optimized to reduce any potential causes of peak broadening.
broadening
The following equation for extra-column broadening shows that it is important to limit injection
Potential causes of peak broadening are:
volume, minimize flow cell volume and make sure that short, narrow ID tubing is used.
Extra-column band broadening
K Constant
Vinj Injection volume The following equation for extra-column broadening shows that it is important to limit injection
volume, minimize flow cell volume and make sure that short, narrow ID tubing is used
Vcell Flow cell volume
F Flow rate
rc Tubing radius
Constant
lc Tubing length Injection volume
Dm Diffusion coefficient in mobileFlow
phasecell volume
Flow rate
Tubing radius
Slow detector response Tubing length Octanophenone
The detector time constant or sampling rate coefficient
Diffusion must be optimized forphase
in mobile narrow peaks. 1 Hz RT: 0.87 – 0.93

If this is not done then losses in intensity and increases in peak width are seen.
Slow detector response 5 Hz

The detector time constant or sampling

Data Peak width rate
Peak Peak height
point* peaks.
must be optimized for narrow 4σ (s)Is this
Area
is (mAu)
10 Hz
1Hzin intensity
not done then losses 2 2.04 increases
and 246330 in 107.4
5Hz
peak width are seen. 6 0.96 57244 118.4
10Hz 10 0.87 55750 114.5
20 Hz
20Hz 18 0.87 55319 115.4
* Number of data points are collected over 4σ 0.87 0.88 0.89 0.90 0.91 0.92 0.93
Minutes

Fast Gradients
For fast gradients it isFast
alsoGradients
important to minimize the pump dwell volume to ensure
that the gradient reaches the column
For fast asitquickly
gradients is also as possible.to minimize
important
the dwell volume to ensure that the gradient
reaches the column as quickly as possible.
Pump dwell volume Column: Hypersil GOLD 1.9µm,
50 x 2.1mm
800µL 80µL
Mobile phase: A – Water + 0.1% Formic Acid
arrives 1.45 min

B – Acetonitrile
arrives 0.15 min

+ 0.1% Formic Acid

Gradient

Gradient: 5 to 100% B in 2 min

Gradient

Flow rate: 0.55mL/min

Temperature: 25°C
Detection: UV@ 270nm (2µL flow cell)
Tubing column – detector: 0.005" ID
0.0 1.0 2.0 3.0 4.0 5.0 0.0 1.0 2.0 3.0 4.0 5.0 Injection volume: 0.5µL
Minutes Minutes
Accucore Technical Guide
Analytes: 1. Sulphaguanidine
2. Sulphamerazine
3. Sulphamonomethoxine
4. Sulphaquinoxaline

14 Thermo Scientific Accucore HPLC Columns | Technical Guide

Reproducible Chromatography
The advanced bonding technology and automated packing process used for Accucore
HPLC columns result in exceptionally reproducible chromatography.

Packing Material Analysis

Every batch of Accucore packing material manufactured is analyzed, compared with specifications
and issued with a certificate of analysis.

Material Properties and Analysis Based on Tanaka Tests

Silica Support Bonded Phase Hydrophobic Secondary Acidic HILIC
Properties Properties Interactions Interactions Interactions Interactions
Hydrophobic HILIC retention
Surface Area Carbon load retention Base activity Acid interaction & selectivity

HR BA AI
Hydrophobic Ion exchange
Pore Size selectivity Chelation capacity pH 2.6

HS C IEX(2.6)
Ion exchange
Particle Size Steric selectivity capacity (pH 7.6)

SS IEX(7.6)
Particle Size Hydrogen bonding
Distribution capacity

HBC

Column Testing
Every Accucore HPLC column is also tested after packing and again compared to specifications
and issued with a certificate.

Retention time Capacity factor Efficiency Asymmetry Backpressure

This material and column testing is evidence that every Accucore HPLC column will
give reproducible chromatography, batch after batch, column after column.

Thermo Scientific Accucore HPLC Columns | Technical Guide 15

 Long Lasting Columns
Chromatographers today demand long lifetimes from the columns they use.

Mechanical Stability and Stable Bonded Phase

The highly uniform packed bed in Accucore HPLC columns is created by the use of tightly controlled
particle size and automated packing process and has excellent mechanical stability.
The advanced bonding technology used for Accucore HPLC columns creates robust bonded phases that
are highly resistant to the effects of pH and temperature.

Accucore HPLC columns show excellent stability at pH <2

280
Column: Accucore C18 2.6µm, 100 x 2.1mm
240
Mobile phase: A – Water + 0.1% Trifluoroacetic Acid
200
Injection 400 B – Methanol + 0.1% Trifluoroacetic Acid
Gradient: Time (min) %B
160 0 25
mAU

0.75 25
120 Injection 200 10.00 100
80
12.00 100
12.20 25
40 17.00 25
Injection 10 Flow rate: 400µL/min
0
0 1 2 3 4 5 6 7 8 9 10 11 Injection volume: 1µL
Minutes Temperature: 30°C
40 Detection: UV at 254nm (0.1s rise time, 20Hz)
35 Order of elution: 1. Uracil (t0)
30 2. Acetaminophen
Capacity Factor

25 3. p-Hydroxybenzoic acid
4. O-Hydroxybenzoic acid
20 5. Amitriptyline
15 6. Nortriptyline
10 7. Di-isopropyl phthalate
8. Di-n-propyl phthalate
5
0
5000 10000 15000 20000 25000 30000 35000
Column Volumes

Accucore HPLC columns

are robust and long lasting.

16 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Accucore HPLC columns are also stable at pH >10
220
Column: Accucore C18 2.6µm, 100 x 2.1mm
Mobile phase: A – Water + 0.1% Ammonia
180 B – Methanol + 0.1% Ammonia
Injection 400
Gradient: Time (min) %B
140 0 15
1.00 15
mAU

8.00 100
100
11.00 100
Injection 220
11.20 15
60 16.00 15
Flow rate: 400µL/min
20 Injection volume: 1µL
Injection 90
0 Temperature: 30°C
0 1 2 3 4 5 6 7 8 9 10 11 Detection: UV at 254nm (0.1s rise time, 20Hz)
Minutes Order of elution: 1. Uracil (t0)
50 2. 4-Chlorocinnamic acid
45 3. Procainamite
4. 4-Pentylbenzoic Acid
40
5. N-Acetylprocainamide
35 6. Di-isopropyl phthalate
Capacity Factor

30 7. Di-n-propyl phthalate
25
20
15
10
5
0
5000 10000 15000 20000 25000 30000 35000
Column Volumes

And also stable at elevated temperature

150
Column: Accucore C18 2.6µm, 100 x 2.1mm
Mobile phase: 35:65 (v/v) Water/Methanol
Flow rate: 400µL/min
Injection 400 Injection volume: 1.5µL
100
Temperature: 70°C
mAU

Detection: UV at 254nm (0.1s rise time, 20Hz)

Injection 130 Order of elution:
50 1. Theophylline/Caffeine (t0)
2. Phenol
3. Butylbenzene
4. o-Terphenyl
Injection 20 5. Pentylbenzene/Triphenylene
0
0 0.2 0.6 1 1.4 1.8 2.2 2.6 3
Minutes

9
8
7
Capacity Factor

6
5
4
3
2
1
0
2000 4000 6000 8000 10000
Column Volumes

Thermo Scientific Accucore HPLC Columns | Technical Guide 17

 Column Protection

Guard columns are designed to protect your column from particulates introduced from the
matrix or instrument and from any strongly retained components in the injected sample.
Thermo Scientific Defender Guard Cartridges have been designed specifically to work
with high speed, high efficiency separations.

Backpressure v Injection Number

1000
Accucore C18 2.6μm, 100mm x 2.1mm
900 With Defender Guard 10mm x 2.1mm Defender Guard Cartridge
1000 Without Guard
800 Mobile: 35:65 (v/v) Acetonitrile/Water
New
With Defender
Defender Guard
Guard
900
700 Without Guard Flow: 550μL/min
(Bar) (Bar)

800 New Defender Guard Temperature: 40°C

600
700 Detection: UV at 254nm
Backpressure

500
600 Injection volume: 1µL
400
Backpressure

500 Protein crash sample: rat plasma diluted 1 in 4 with Acetonitrile

300
400 1 injection test mix followed by 9 protein crash injections
200
300
100
200
0
100 0 100 200 300 400 500 600 700 800

0
Injection Number Defender Guard Cartridges extend the
0 100 200 300 400 500
Injection Number
600 700 800
lifetime of your Accucore HPLC column.

120

100 Accucore C18 2.6μm, 100mm x 2.1mm

120 10mm x 2.1mm Defender Guard Cartridge
80
(MilliVolts)

100 Mobile phase: 50:50 (v/v) Acetonitrile/Water

60 Flow rate: 400μL/min
80 With Defender
(MilliVolts)

40 Guard Cartridge Temperature: 30°C

60
Response

With Defender Detection: UV at 254nm

20 Guard Cartridge
40 Injection volume: 1μL
Response

0
20 Analytes: 1. Theophylline
-20 2. p-Nitroaniline
0 3. Methyl Benzoate
Without Guard
-40 4. Phenetole
-20
5. o-Xylene
-60 Without Guard
-40
0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5
-60
Minutes
0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5
Minutes

With Without Defender Guard Cartridges have

Parameter Defender Guard Defender Guard minimal effect on column performance.
Selectivity 1.81 1.81
Efficiency (N/m) 185672 213350
Tailing 1.18 1.07
Retention Time (minutes) 3.19 2.90
Backpressure (bar) 280 256

18 Thermo Scientific Accucore HPLC Columns | Technical Guide

Optimum Selectivity
Accucore HPLC columns are available in a series of phases to provide a wide range of selectivities.
Each of the bonded phases is manufactured using advanced bonding technology and is characterized
using a testing regime based on the Tanaka Tests. See page 15 for further details of these tests.

The radar plots below show the results of the characterization and allow for quick and easy
comparison of the phase selectivities.

Accucore RP-MS
Optimized for MS detection, excellent combination of speed
and quality of separation

Accucore C18
Optimum retention for non-polar analytes

Accucore aQ
Compatible with 100% aqueous mobile phases, special selectivity
for polar analytes

Phenyl-Hexyl
Unique selectivity for aromatic and moderately polar analytes

Accucore PFP
Alternative selectivity to C18, particularly for halogenated analytes

Accucore HILIC
HILIC
Enhanced retention of polar and hydrophilic analytes

Thermo Scientific Accucore HPLC Columns | Technical Guide 19

 Thermo Scientific Accucore RP-MS
Hydrophobicity

Low High
pH Range

0 2 to 9 14
Pore Size

0 80Å 300
Carbon Load (%)

0 7 25

Particle Size 2.6µm

• Optimized for MS detection

• Excellent Peak Shapes
• Excellent Combination of Speed and Efficiency
Accucore RP-MS uses an optimized alkyl chain length for more effective coverage of the silica surface. This coverage
results in a significant reduction in non-hydrophobic interactions and thus highly efficient peaks with very low tailing.
RP-MS offers slightly lower retention than C18 and this combined with high efficiencies and low peak tailing make
this the phase of choice for use with MS detection.
The selectivity offered by Accucore RP-MS matches that of C18 columns.

Bases

240 Accucore RP-MS 2.6µm, 50mm x 2.1mm

4
220 Mobile phase: 65:35 (v/v) Methanol/25mM
200 2 Potassium Phosphate pH7.0
180 Flow rate: 500µL/min

160
Temperature: 30°C
Detection: UV at 215nm
140
1 Injection volume: 1µL
mAU

120
Back pressure: 232 bar
100
3 Analytes: 1.Uracil
5
80 2. Propranolol
6
60 3. Butylparaben
4. Naphthalene
40
5. Acenaphthene
Tf = 1.27
20 6. Amitriptyline
0
0.0 0.5 1.0 1.5 2.0 2.5 3.0
Minutes

20 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Thermo Scientific Accucore C18
Hydrophobicity

Low High
pH Range

0 1 to 11 14
Pore Size

0 80Å 300
Carbon Load (%)

0 9 25

Particle Size 2.6µm USP L1

• Optimum retention of non-polar compounds

• Hydrophobic interaction mechanism
• Separates a broad range of analytes
The carbon loading of Accucore C18 phase provides high retention of non-polar analytes via a predominantly
hydrophobic interaction mechanism.
The highly retentive nature of Accucore C18 phase means that it can be used to separate a broad range of analytes.

Triazines

100 Accucore C18 2.6µm, 50mm x 2.1mm

90 3 Mobile phase: A – Water ; B – Acetonitrile
80 Gradient: Time (min) %B
2 1.0 35
70
4 2.5 70
6
60 Flow rate: 600µL/min
mAU

1
50 Temperature: 25°C
40 Detection: UV at 280nm
5
30 Injection volume: 2µL
20 Backpressure: 298 bar
10 Analytes: 1. Simazine
2. Simetryn
0 3. Atrazine
0.0 0.5 1.0 1.5 2.0 2.5 3.0 4. Ametryn
Minutes 5. Propazine
6. Prometryn

Thermo Scientific Accucore HPLC Columns | Technical Guide 21

 Thermo Scientific Accucore aQ
Hydrophobicity

Low High
pH Range

0 2 to 9 14
Pore Size

0 80Å 300
Carbon Load (%)

0 9 25

Particle Size 2.6µm USP L1

• Retention and Resolution of Polar Analytes

• Polar Endcapped C18 Stationary Phase for Alternative Selectivity
• Ideal for Highly Aqueous Mobile Phases
The polar functional group used to endcap Accucore aQ phase provides an additional controlled interaction
mechanism by which polar compounds can be retained and resolved, making Accucore aQ phase ideal for the
quantitative analysis of trace levels of polar analytes.
The wettability of reversed phase media can be increased by the introduction of polar functional groups. The polar
endcapping of Accucore aQ media also makes it usable in 100% aqueous mobile phases without the risk of loss
of performance or poor stability.

Lamivudine (USP)
Accucore aQ 2.6µm, 50mm x 2.1mm
140
Mobile phase: 95:5 (v/v) Ammonium Acetate,
120 pH 3.80 / Methanol
Flow rate: 200µL/min
100
Temperature: 35°C
mAU

80 Detection: UV at 277nm
60 Injection volume: 1µL
Analytes: Lamivudine
40 %RSD Tr 0.00
%RSD Peak area 1.72
20
(%RSD calculated from 6 replicate injections)
0 USP acceptance criteria: % RSD (Tr, Peak Area) <2.0
0.0 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 3.0
Minutes

22 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Thermo Scientific Accucore Phenyl-Hexyl
Hydrophobicity

Low High
pH Range

0 2 to 8 14
Pore Size

0 80Å 300
Carbon Load (%)

0 5 25

Particle Size 2.6µm USP L11

• Mixed-mode Selectivity for Aromatic and Moderately Polar Analytes

• Enhanced Pi-pi Interactions with Aromatics
• Moderate Hydrophobicity
The C6 chain in Accucore Phenyl-Hexyl phase exhibits classical RP retention and selectivity, while the phenyl ring can add
special selectivity by interacting with polar groups within the solutes. This results in a mixed-mode separation mechanism.
Phenyl-Hexyl phase should be selected for complex samples where some peaks are well resolved on a conventional
alkyl phases, but are not well resolved on a conventional phenyl phase. While other peaks are well resolved on a phenyl
phase, but not well resolved on a conventional alkyl phase.

Beta-agonists

100 2.89 Accucore Phenyl-Hexyl 2.6µm, 100mm x 2.1mm

NL: 9.22E5 Mobile phase: A – Ammonium acetate 5mM, pH 4
Terbutaline m/z = 225.60-226.60
0 B – Acetonitrile
100 3.04 Gradient: Time (min) %B
NL: 6.51E5 0 5
Cimaterol 1 5
0 m/z = 239.60-240.60 MS
100 3.16 10 100

NL: 3.52E6 Flow rate: 0.25mL/min

Salbutamol Temperature: 40°C
0 m/z = 219.60-220.60
100 4.72 Detection: +ESI-MS (45ºC, 4.5kV, 60V, scan 150 – 350)
NL: 2.23E6 Injection volume: 1µL
Ractopamine
0 m/z = 301.60-302.60 MS Backpressure: 120 bar
100 4.80
NL: 2.51E6
Clorprenaline
0 m/z = 213.60-214.60 MS
100 5.13
NL: 1.71E6
Clenbuterol
0 m/z = 276.60-277.60 MS
0 1 2 3 4 5 6 7
Minutes

Thermo Scientific Accucore HPLC Columns | Technical Guide 23

 Thermo Scientific Accucore PFP
Hydrophobicity

Low High
pH Range

0 2 to 8 14
Pore Size

0 80Å 300
Carbon Load (%)

0 5 25

Particle Size 2.6µm USP L43

• Alternative Selectivity to C18

• Extra Retention for Halogenated Species
• Unique Selectivity for Non-halogenated Polar Compounds
Introduction of fluorine groups into the Accucore PFP (pentafluorophenyl) stationary phase causes significant changes
in solute-stationary phase interactions. This can lead to extra retention and selectivity for positional isomers of
halogenated compounds.
PFP Columns are also well suited to the selective analysis of non-halogenated compounds, in particular polar
compounds containing hydroxyl, carboxyl, nitro, or other polar groups. High selectivity is often most apparent when
the functional groups are located on an aromatic or other rigid ring system.

Positional Isomers

70 Accucore PFP 2.6µm, 50mm x 2.1mm

Mobile phase: A – Water + 0.1% Formic Acid
60 B – Acetonitrile + 0.1% Formic Acid
Gradient: 15 – 30% B in 7 minutes
50
5
Flow rate: 600µL/min

40
Temperature: 50°C
7 Detection: UV at 270nm
mAU

30 1 Injection volume: 2µL

8 9 1. 3,4 – Dimethoxyphenol
10
20 2 2. 2,6 – Dimethoxyphenol
11 12
13 14 3. 2,6 – Difluorophenol
10 34 6 4. 3,5 – Dimethoxyphenol
5. 2,4 – Difluorophenol
0
6. 2,3 – Difluorophenol
7. 3,4 – Difluorophenol
8. 3,5 – Dimethylphenol
0 1 2 3 4 5 6 7 8
9. 2,6 – Dimethylphenol
Minutes 10. 2,6 – Dichlorophenol
11. 4 – Chloro-3-Methylphenol
12. 4 – Chloro-2-Methylphenol
13. 3,4 – Dichlorophenol
14. 3,5 – Dichlorophenol

24 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Thermo Scientific Accucore HILIC
Hydrophobicity

Low Very Low (5%) High

HILIC
pH Range

0 2 to 8 14
Pore Size

0 80Å 300

Particle Size 2.6µm USP L3

• Enhanced Retention of Polar and Hydrophilic Analytes

• Alternative Selectivity to C18 without Ion-pair or Derivatization
• Improved Sensitivity for MS detection
Analyte properties that govern retention with Accucore HILIC phase are acidity/basicity, which determines hydrogen
bonding, and polarizability which determines dipole-dipole interactions.
The highly organic mobile phases used with Accucore HILIC phase ensure efficient desolvation in ESI MS detection,
which in turn leads to improved sensitivity.

Catecholamines
2 Accucore HLIC 2.6µm, 150 x 4.6mm
140 Mobile phase: 85:15 (v/v) Acetonitrile/100mM
Ammonium Formate, pH 3.2
120 Flow rate: 2mL/min
1 Temperature: 40°C
100 4
Detection: UV at 280nm
80 Injection volume: 5µL
mAU

3
Backpressure: 157 bar
60 Analytes: 1. Catechol
2. 5-HIAA
40
5 3. DOPAC
6
4. Serotonin
20 7 5. L-tyrosine
6. Dopamine
0 7. L-DOPA
0 0.5 1 1.5 2 2.5 3 3.5 4
Minutes

Thermo Scientific Accucore HPLC Columns | Technical Guide 25

 Applications
Food
Azo Dyes

70 Accucore RP-MS 2.6µm, 150mm x 4.6mm

60
Mobile phase: A – Water + 0.1% Trifluoroacetic Acid
B – Acetonitrile + 0.1% Trifluoroacetic Acid
50 Gradient: Time (min) %B
mAU

40 7
0 25
6
5 3 30
4
30 15 95
2 3 17 95
20
Flow rate: 1.8mL/min
10 1
Temperature: 50°C
0 Detection: UV at 225nm
-100 Injection volume: 2µL
0 2 4 6 8 10 12 14 16
Analytes: 1. Fast Garnet
Minutes 2. Orange II
3. Dimethyl Yellow
4. Sudan Red G
5. Sudan I
6. Sudan II
7. Sudan IV

Melamine and Cyanuric Acid

Accucore HILIC 2.6µm, 150mm x 4.6mm

2
100 m/z = 126.50- 128.20 Mobile phase: 90:10 (v/v) Acetonitrile:50mM
167.50- 168.50 Ammonium Acetate, pH 5
80 Flow rate: 1mL/min
mAU

Temperature: 40°C
60 Detection: MS at m/z 127, 128, 168
(negative mode 0-3 mins,
40 positive mode 3-10 mins)
Injection volume: 5µL
20 1 Backpressure: 117 bar
Analytes: Cyanuric Acid: m/z 128.1 (-1)
0 Melamine: m/z 127.1 (+1),
0 1 2 3 4 5 6 7 8 9 168.1 (+1 with Acetonitrile adduct)
Minutes

26 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Environmental
Uron Herbicides

3 Accucore RP-MS 2.6µm, 100mm x 2.1mm

4
60000
Mobile phase: A – Water; B – Acetonitrile
48000 2 Gradient: 35 – 60% B in 3.20 min
Flow rate: 0.43mL/min
mAU

36000 1
5 6 Temperature: 25°C
24000 Detection: UV at 240nm
Injection volume: 3µL
12000 Analytes: 1. Monuron
2. Metoxuron
0 3. Linuron
0 1 1.5 2 2.5 3 3.5 4 4. Diuron
Minutes 5. Terbuthiuron
6. Chlortoluron

Banned Aromatic Amines

Accucore PFP 2.6µm, 100mm x 2.1mm

240 4
Mobile phase: A – 25mM Ammonium Acetate pH 5.0
B – Acetonitrile
200
Gradient: 20 – 100% B in 1.5 minutes
160 Flow rate: 0.6mL/min
3
Temperature: 40°C
mAU

120 1
2 6 Detection: UV at 254nm
Injection volume: 1µL
80
Backpressure: 300 bar
5
40 Analytes: 1. 2,4 – Diaminotoluene
2. 4,4 – Oxydianiline
3. o-Toluidine
0
4. 2-Methyoxy-5-methylaniline
0 0.2 0.4 0.6 0.8 1 1.2 1.4 2
5. 2,4,5-Trimethylaniline
Minutes 6. 4,4-Methylene-bis (2-chloroaniline)

Basic polar compounds, volatile buffer

in the mobile phase – good peak shape.

Thermo Scientific Accucore HPLC Columns | Technical Guide 27

 Clinical

Epinephrine and Norepinephrine

60 Accucore HILIC 2.6µm, 150mm x 4.6mm
Mobile phase: 92:8 (v/v) Acetonitrile/100mM
50 Ammonium Formate, pH 3.2
Flow rate: 3.5mL/min
40 Temperature: 40°C
Detection: UV at 230nm
30 Injection volume: 10µL
mAU

Backpressure: 250 bar

20
Analytes: 1. Epinephrine
2. Norepinephrine
10

0
-5
0 1 2 3 4 5 6
Minutes

1
350 2 Accucore aQ 2.6µm, 100mm x 2.1mm

300
Mobile phase: 5mM Ammonium Formate, pH3.2
Flow rate: 0.6mL/min
250 Temperature: 30°C
Detection: UV at 230nm
200
Injection volume: 2µL
150 Analytes: 1. Norepinephrine (0.33 min)
mAU

2. Epinephrine (0.37 min)

100

50

0 0.5 1 1.5 2
Minutes

Good retention and resolution of the two analytes in HILIC

compared with the aQ column in reversed phase mode.

28 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Steroids

90 1 Accucore RP-MS 2.6µm, 100mm x 2.1mm

2 Mobile phase: 50:50 (v/v) Water / Acetonitrile
80
Flow rate: 0.6mL/min
70
Temperature: 40°C
60
Detection: UV at 220nm
50
Injection volume: 1µL
40
mAU

Analytes: 1. Estradiol
30 2. Estrone
20

10

0 0.25 0.5 0.75 1.0 1.25 1.5

Minutes

Speed – Sub 1 minute separation.

35 Accucore RP-MS 2.6µm, 100mm x 2.1mm

4
5 Mobile phase: A – Water; B – Acetonitrile
30 3
1 Gradient: 25 – 70% B in 4 minutes
25 2 6 Flow rate: 0.6mL/min
Temperature: 25°C
20
Detection: UV at 254nm
15 Injection volume: 1µL
mAU

Analytes: 1. Hydrocortisone
10 2. Cortisone
3. Corticosterone
5 4. 11-alpha Hydroxyprogesterone
5. 17-alpha Hydroxyprogesterone
0 6. Progesterone

0 1 2 3 4 5 6
Minutes

Thermo Scientific Accucore HPLC Columns | Technical Guide 29

 Forensic Toxicology
Testosterones
2
600 Accucore RP-MS 2.6µm, 100mm x 2.1mm
1
Mobile phase: 60:40 (v/v) Water / Acetonitrile
500 3
Flow rate: 0.6mL/min
400 Temperature: 40°C
Detection: UV at 254nm
300 Injection volume: 1µL
mAU

Analytes: 1. 11-Ketotestosterone
200
2. 19-Nortestosterone (Nandrolone)
3. Epitestosterone
100

0 0.5 1 1.5 2 2.5

Minutes

Forensics
Explosives

2
30

25

20 3
1
mAU

15
5
10 Accucore PFP 2.6µm, 100mm x 2.1mm
4
Accucore C18 2.6µm, 100mm x 2.1mm
5
PFP Mobile phase: 60:40 (v/v) Water/Methanol
0 Flow rate: 0.4mL/min
Temperature: 40°C
0 2 4 6 8 10
Detection: UV 254nm
Minutes
100 Injection volume: 1µL
3 Backpressure: 260 bar
80 Analytes: 1. 1,3,5-trinitrobenzene
2. Nitrobenzene
1 3. 1,3-dinitrobenzene
60 5
4. 2,4,6-trinitrotoluene
5. 2,4-dinitrotoluene
mAU

2
40

4
20

C18
0

0 2 4 6 8 10
Minutes

The carbon-fluorine bond on the PFP bonded phase is more polar than the
carbon-hydrogen bond in the C18 phase, which results in extra retention and
selectivity of these polar substituted aromatic compounds containing nitro
functional groups.

30 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Pharmaceuticals
Acidic and Neutral Drugs

1 Accucore C18 2.6µm, 100mm x 2.1mm

30
6 Mobile phase: A – Water + 0.1% Phosphoric Acid
4
25 B – Acetonitrile + 0.1% Phosphoric Acid
20 5 Gradient: Min %B
mAU

0 30
15 7 2 40
2 4 95
10 3
Flow rate: 0.4mL/min
5
Temperature: 25°C
0 Detection: UV at 215nm
Injection volume: 1.5µL
0 1 2 3 4 5 6 7
Minutes
Analytes: 1. Hydrochlorothiazide
2. Prednisolone
3. Pravastatin
4. Carbamazepine
5. Diclofenac
6. Ibuprofen
7. Progesterone

Excellent resolution.

Zidovudine (USP)

350
1 Accucore C18 2.6µm, 100mm x 2.1mm
300 Mobile phase: 80:20 (v/v) Water/Methanol
Flow rate: 0.5mL/min
250
Temperature: 25°C
200 Detection: UV at 265nm
mAU

150 Injection volume: 1µL

Analytes: 1. Zidovudine USP specification
100
Asymmetry 1.13 < 2.0
50 %RSD tr 0.0 < 2.0
%RSD Peak area 0.34 < 2.0
0

0 0.5 1 1.5 2 2.5 3

Minutes

The Accucore C18 column demonstrated

excellent performance for the analysis of
Zidovudine. The USP criteria was exceeded.

Thermo Scientific Accucore HPLC Columns | Technical Guide 31

 Ibuprofen

80 Accucore C18 2.6µm, 100mm x 4.6mm

2
70 Mobile phase: 66.3% Water adjusted to pH 2.5 with
60 Phosphoric Acid / 33.7% Acetonitrile
50 Flow rate: 2.0mL/min
mAU

40 Temperature: 30°C
30 Detection: UV at 214nm
20 Injection volume: 5µL
1
10 Back pressure: 276 bar
0 Analytes: 1. Valerophenone (USP grade)
2. Ibuprofen (USP grade)
0 2 6 10 14 18 22 26 30 34
Minutes

The Accucore C18 column demonstrated excellent performance

for the analysis of Ibuprofen, using a method based on the USP.

Fenoprofen and Gemfibrozil (USP)

1 Accucore RP-MS 2.6µm, 100mm x 2.1mm

40 Mobile phase: 50:49.6:0.4 (v/v/v)
35 Acetonitrile/Water/Phosphoric Acid
30 Flow rate: 0.4mL/min
25 Temperature: 30°C
mAU

20 Detection: UV at 272nm
15 Injection volume: 3µL
2
10 Analytes: 1. Fenoprofen 2. Gemfibrozil USP
specification
5
Asymmetry 1.23 1.22 <2
0 %RSD tr 0.39 0.39 <2
%RSD Peak area 0.39 0.00 <2
0 1 2 3 4 5 Resolution > 8
Minutes Efficiency 7464 8259 > 3000

32 Thermo Scientific Accucore HPLC Columns | Technical Guide

 Lisinopril

80 3 Accucore RP-MS 2.6µm, 100mm x 2.1mm

70 Mobile Phase: A – 0.026M Monobasic Sodium
60 2 Phosphate pH 3.75 with Phosphoric Acid
B – 80:20 (v/v) Mobile Phase A/Acetonitrile
50
Gradient: Time (min) %B
0 3
mAU

40
15 30
30 30 30
20 35 3
4
Flow rate: 0.4mL/min
10
1 Temperature: 50°C
0
Detection: UV at 210nm
0 5 10 15 20 25 30 Injection volume: 5µL
Minutes Backpressure: 159 bar
Analytes: 1. Impurity 1
2. 2-amino-4-Phenylbutyric Acid
3. Lisinopril
4. Lisinopril R,S.S isomer
Analytes: Lisinopril USP specification
Resolution 2,1 4.7 > 1.5
Resolution 2,3 12.7 > 1.5
Resolution 4,3 4.7 > 2.0
%RSD Peak 1.74 < 5%
Area Lisinopril

The Accucore RP-MS column demonstrated excellent performance

for the analysis of Lisinopril. The USP criteria was exceeded.

Bio-pharmaceuticals
Peptide Test Mix

1 Accucore C18 2.6µm, 100mm x 2.1mm

300 Mobile phase: A – Water + 0.1% Trifluoroacetic Acid
B – Acetonitrile + 0.1% Trifluoroacetic Acid
250 3 6
2 Gradient: 10 – 70 % B in 6 min
200 Flow rate: 0.50mL/min
150 4
Temperature: 40°C
Detection: UV at 220nm
mAU

100
5 Injection volume: 2µL
50 Analytes: 1. Glycyl-L-Tyrosine (MW=238.3)
2. Val-Tyr-Val (MW=379.5)
0
3. Methionine Enkephalin (MW=573.7)
-50 4. Leucine Enkephalin (MW=555.6)
5. Ribonuclease A (MW=13700)
6. Insulin (MW=5733.5)
0 1 1.5 2 2.5 3 3.5 4
Minutes

The Accucore C18 column can be used to analyze small peptides;

demonstrated up to MW 13700.

Thermo Scientific Accucore HPLC Columns | Technical Guide 33

 General

High Throughput Ketones

2 Accucore C18 2.6µm, 50mm x 2.1mm

160000
Mobile phase: A – Water; B – Acetonitrile
140000
Gradient: Min %B
120000 0 40
4
100000 0.4 95
mAU

80000 5 0.80 95
1 2 Flow rate: 1.0mL /min
60000
160000 6
7 Temperature: 45°C
140000
40000 3
Detection: UV at λ = 258nm
120000
20000
Injection volume: 1µL
4
1000000 Backpressure: 113 bar
mAU

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 Analytes: 1. 2-Pentanone
80000 5
Minutes 2. Acetophenone
60000 1
6 3. 2-Heptanone
3 7 4. Butyrophenone
40000
5. Hexanophenone
20000 6. Octanophenne
600
2
7. Decanophenone
0
500 0 0.1 0.2 0.3 0.4
4
0.5 0.6 0.7 0.8 0.9
3 Minutes 6
400 1 5 7
mAU

300

600 Hypersil GOLD C18 5µm, 150mm x 4.6mm

200 2
Mobile phase: A – Water; B – Acetonitrile
500 4 Gradient: Min %B
100
3 6 0.00 40
400 1 5 7 2.30 95
0
4.60 95
mAU

0 2.0 2.4 2.8 3.2 3.6 4.0 4.4 4.8 5.2 5.6
300 Flow rate: 1.0 mL/min
Minutes
Temperature: 45°C
200
Detection: UV at λ = 258nm
100 Injection volume: 10µL
Pressure: 47 bar
0
Analytes: 1. 2-Pentanone
0 2.0 2.4 2.8 3.2 3.6 4.0 4.4 4.8 5.2 5.6
2. Acetophenone
Minutes 3. 2-Heptanone
4. Butyrophenone
5. Hexanophenone
6. Octanophenne
7. Decanophenone

Separation with the Accucore HPLC column is 6 times

faster than the 150 x 4.6mm, 5µm Hypersil GOLD column.

34 Thermo Scientific Accucore HPLC Columns | Technical Guide

Thermo Scientific Options for Fast HPLC
Uniquely in the market Thermo Scientific HPLC columns offer chromatographers
three different options for fast HPLC.

Hypersil GOLD 1.9µm and Syncronis 1.7µm – Small Particle Advantage

When used with UHPLC systems Hypersil GOLD 1.9µm and Syncronis 1.7µm HPLC columns offer
optimum efficiency over a range of flow rates.
This high efficiency results in higher throughput, lower costs and better separations.

Accucore – Core Enhanced Technology

The core enhanced technology used in Accucore HPLC columns provides better speed, sensitivity
and resolving power than 5µm and 3µm columns without the higher backpressures associated with
sub-2µm materials.
Accucore HPLC columns do not require UHPLC systems.

Hypercarb – High Temperature HPLC

Hypercarb is a porous graphitized carbon material suitable for use at temperatures up to 200°C.
As a result it is uniquely suited for use with high temperature HPLC.
High temperatures in HPLC increase efficiency, reduce backpressure and reduce analysis time.

Thermo Scientific Accucore HPLC Columns | Technical Guide 35

 Ordering Information

Accucore HPLC Columns

Description Particle Size Length (mm) 2.1mm ID 3.0mm ID 4.6mm ID
30 17626-032130 17626-033030 17626-034630
50 17626-052130 17626-053030 17626-054630
Accucore RP-MS 2.6µm
100 17626-102130 17626-103030 17626-104630
150 17626-152130 17626-153030 17626-154630
30 17126-032130 17126-033030 17126-034630
50 17126-052130 17126-053030 17126-054630
Accucore C18 2.6µm
100 17126-102130 17126-103030 17126-104630
150 17126-152130 17126-153030 17126-154630
30 17326-032130 17326-033030 17326-034630
50 17326-052130 17326-053030 17326-054630
Accucore aQ 2.6µm
100 17326-102130 17326-103030 17326-104630
150 17326-152130 17326-153030 17326-154630
30 17926-032130 17926-033030 17926-034630
50 17926-052130 17926-053030 17926-054630
Accucore Phenyl-Hexyl 2.6µm
100 17926-102130 17926-103030 17926-104630
150 17926-152130 17926-153030 17926-154630
30 17426-032130 17426-033030 17426-034630
50 17426-052130 17426-053030 17426-054630
Accucore PFP 2.6µm
100 17426-102130 17426-103030 17426-104630
150 17426-152130 17426-153030 17426-154630
30 17526-032130 17526-033030 17526-034630
50 17526-052130 17526-053030 17526-054630
Accucore HILIC 2.6µm
100 17526-102130 17526-103030 17526-104630
150 17526-152130 17526-153030 17526-154630

Accucore Defender Guard Columns (4/pk)

Description Particle Size Length (mm) 2.1mm ID 3.0mm ID 4.6mm ID
Accucore RP-MS 2.6µm 10 17626-012105 17626-013005 17626-014005
Accucore C18 2.6µm 10 17126-012105 17126-013005 17126-014005
Accucore aQ 2.6µm 10 17326-012105 17326-013005 17326-014005
Accucore Phenyl-Hexyl 2.6µm 10 17926-012105 17926-013005 17926-014005
Accucore PFP 2.6µm 10 17426-012105 17426-013005 17426-014005
Accucore HILIC 2.6µm 10 17526-012105 17526-013001 17526-014005

UNIGUARD Direct-Connection Guard Cartridge Holders

Description 2.1mm ID 3.0mm ID 4.6mm ID
UNIGUARD Drop-In Guard Cartridge Holder 852-00 852-00 850-00
Standard Replacement Tip 850-RT 850-RT 850-RT

36 Thermo Scientific Accucore HPLC Columns | Technical Guide

Accucore Kits
Accucore kits allow validation of the reproducibility of Accucore HPLC columns,
or verification of which Accucore HPLC column offers the best separation.

Accucore Validation Kit

Validate the reproducibility of Accucore HPLC columns. Contains 3 Accucore C18 columns.
Description Particle Size Length (mm) 2.1mm ID
Accucore 2.6µm 50 17126-052130-3V
Validation Kit 100 17126-102130-3V
150 17126-152130-3V

Accucore Narrow Selectivity Kit

Verify which Accucore HPLC column offers the best separation over a narrow selectivity range.
Contains 1 each of Accucore C18, RP-MS and aQ columns.
Description Particle Size Length (mm) 2.1mm ID
Accucore Narrow 2.6µm 50 17X26-052130-3VA
Selectivity Kit 100 17X26-102130-3VA
150 17X26-152130-3VA

Accucore Wide Selectivity Kit

Verify which Accucore HPLC column offers the best separation over a wide selectivity range.
Contains 1 each of Accucore C18, Phenyl-Hexyl and PFP columns.
Description Particle Size Length (mm) 2.1mm ID
Accucore Wide 2.6µm 50 17X26-052130-3VB
Selectivity Kit 100 17X26-102130-3VB
150 17X26-152130-3VB

Accucore Polar Selectivity Kit

Verify which Accucore HPLC column offers the best separation for polar analytes.
Contains 1 each of Accucore aQ, PFP and HILIC columns.
Description Particle Size Length (mm) 2.1mm ID
Accucore Polar 2.6µm 50 17X26-052130-3VC
Selectivity Kit 100 17X26-102130-3VC
150 17X26-152130-3VC

Thermo Scientific Accucore HPLC Columns | Technical Guide 37

 Resources
for Chromatographers

Thermo Scientific Chromatography

Columns and Consumables Catalog
This extensive catalog offers 540 pages of proven
chromatography tools and product selection guides.
Available online, with a robust search tool and
optimized for your iPad®.
Visit www.thermoscientific.com/catalog

Chromatography Resource Center

Our web-based resource center provides technical support,
applications, technical tips and literature to help move your
separations forward.
Visit www.thermoscientific.com/chromatography

For more information visit: www.thermoscientific.com/accucore

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