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Examples of transfer from a conventional 5
µ
m column to SunShell column
Isocratic separation
0
1
2
3
4
5
6
Retention time/min
0
1
2
3
4
5
6
Retention time/min
0 2
4
6
8 10 12 14 16 18 20 22 24
Retention time/min
HPLC
UHPLC
1/3 of analysis time
1
1
1
1
1
2
2
2
2
2
3
3
3
3
3
4
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4
4
4
5
5
5
5
5
ACE C18, 5
µ
m 250 x 4.6 mm
SunShell C18, 2.6
µ
m 100 x 4.6 mm
1.0 mL/min
1.8 mL/min
N(4)=20,287
N(4)=22,424
N(4)=19,313
N(4)=24,124
N(4)=20,287
Analysis time decreases to 1/6 to compare
with a 5
µ
m column sized 250 x 4.6 mm.
Column:
ACE C18, 5
µ
m 250 x 4.6 mm
SunShell C18, 2.6
µ
m 100 x 4.6 mm
Mobile phase:
CH
3
CN/20mMPhosphoric acid = 45/55
Flow rate: 1.0 mL/min,
1.8 mL/min at the lowest chromatogram
Temperature: 25
º
C
Pressure: 9.5 MPa for ACE C18 5
µ
m
13.4 MPa for SunShell C18 2.6
µ
m
Detection: UV@230 nm
Sample: 1 = Benzydamine
2 = Ketoprofen
3 = Naproxen
4 = Indomethacin
5 = Ibuprofen
HPLC: Hitachi LaChrom ELITE
(using 0.25 mm i.d. tubing)
UHPLC: Jasco X-LC
Gradient separation
0
1
2
3
4
5
6
Retention time/min
0
1
2
3
4
5
6
Retention time/min
Time 0 min 15 min 20 min
%B 2% 25% 25%
Time 0 min 7.5 min 10 min
%B 2% 25% 25%
0
2
4
6
8
10
12
14
16
18
20
22
24
Retention time/min
HPLC
UHPLC
1/2 of Analysis time
1
1
1
1
2
2
2
2
3
3
3
3
4
4
4
4
5
5
5
5
Sunniest C18, 5
µ
m 150 x 4.6 mm
SunShell C18, 2.6
µ
m 75 x 4.6 mm
7
8
9
6
6
6
7
7
8 9
7
8 9
8 9
8 9
7 6
Column:
Sunniest C18, 5
µ
m 150 x 4.6 mm
SunShell C18, 2.6
µ
m 75 x 4.6 mm
Mobile phase:
A) 0.1% Phosphoric acid
B) CH
3
CN
Gradient program for Sunniest C18
for SunShell C18
Flow rate: 1.0 mL/min,
Temperature: 25
º
C
Detection: UV@250 nm
Sample: Oolong tea
1 = Gallocatechin, 2 = Epigallocatechin,
3 = Catechin, 4 = Caffeine, 5 = Epicatechin,
6 = Epigallocatechin gallate, 7 = Gallocatechin
gallate, 8 = Epicatechin gallate, 9 = Catechin gallate
HPLC: Hitachi LaChrom ELITE
(using 0.25 mm i.d. tubing)
UHPLC: Jasco X-LC
There are difference of system time lag
between HPLC and UHPLC. UHPLC has much
less system time lag than HPLC because of high
pressure gradient system for UHPLC and low
pressure gradient system for HPLC.
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