TR02AE

The times they are a-changin', from 5um to 3um ... Unison series

Aminopropyl type, Normal phase HPLC column

Revolutionary aqueous durability for aminopropyl phase
Aqueous to non-aqueous Normal Phase Separation
3um particle high-speed and superior resolution
LC-MS applicable
Pure spherical porous silica / 3um particle / Aminopropyl phase

Unequalled durability against water elution
2

1. fructose(2.5mg/mL)

120

1sucrose3

2. glucose(3.75mg/mL)

100

4

3. sucrose

80(2.5mg/mL)

Unison UK-Amino, 250 x 2 mm

(3.75mg/mL)

500 hr% tR

4. maltose

60water, 0.1mL/min, ambient
40400 hr

20220 hr

150 hrInitial0510 min00

Conventional Aminopropyl Column

100

200 300hours

400500Analytical Conditions

Unison UK-Amino, 250 x 2 mm
acetonitrile /water = 75 /25
0.2mL/min (8MPa)
37 deg.C, 4uL, ELSD

304

Aqueous durable silica-based aminopropyl columns have been used for a long time as a normal phase column
for carbohydrate separation. However, these aminopropyl columns have a fatal flaw: "column bleeding" or therapid deterioration in retention as a result of ligand desorption under aqueous elution.
Our newly-designed Unison UK-Amino offers a different approach from conventional columns: high durability
against aqueous eluent. As the above chromatogram demonstrates, conventional columns experience asignificant decline in retention when an aqueous mobile phase elutes through the column. UK-Amino, on theother hand, does not show any change in separation or retention. This is a significant development in the historyof aminopropyl columns.
UK-Amino's design not only provides analytical power, but the 3um particle high-resolution column has otherbenefits including the minimization of LC-MS and LC-ELSD noise levels. UK-Amino can be applied to aqueous
normal phases conditions, and separation optimization is possible while comparing to ODS columns using
reversed-phase mode. One can expect significant results from this normal phase column of UK-Amino.
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North America

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Other Countries

TR02AE-JF15

Normal Phase and Anion Exchange Modes
Aminopropyl stationary phases generally employ both a normal phase separation mode andan anion exchange mode derived from amino groups. There are two methods using Unison
UK-Amino to separate ascorbic acid an its isomer isoascorbic acid (erythorbic acid).
316

1

12

HOH

HOCH22CO

O

HO1

21

HOCH22

C

O

HOOH

2

L-ascorbic acid(Vitamin C)

Normal phase +anion exchange
4 chromatograms from the topOHH

On the normal phase mode,
retention deteriorates as thepolarity in the mobile phaserises.
However,astheacetonitrile partition rises,
retention increases and the twocompounds
completelyseparate at 75% acetonitrile.
Moreover, the elution order isreversed from anion exchange
mode due to the difference ininteractions.

OH

isoascorbic acid(erythorbic acid)

Unison UK-Amino, 250 x 2 mm
0.2 mL/min (9-14 MPa)
37 deg.C, 260nm
2.5mg/mL, 2uL

175% ACN

3% AcOH

50% ACN

3% AcOH

20% ACN

3% AcOH

0% ACN

3% AcOH

0% ACN

1% AcOH

0% ACN

0.5% AcOH

12normal phase

Anion exchange
3 chromatogramsfrom the bottom
In anion exchange mode,
retention deteriorates as theionic strength grows larger andpH-driven ionic interactions
grow weaker (in this case, highacidic density = low pH). In thisexample, two compounds arecompleted separated with only
a 0.5% acetic acid aqueoussolution.

1 2anion exchange

01020 minNormal Phase Separation of Saccharides
Unison UK-Amino provides excellent peak shape with 3um particle for hydrophilicmonosaccharides and sugar alcohols separation.
Monosaccharides

Sugar Alcohols

305

2

1O

HO
CH33OH

meso-erythritol

OHO
OH

CH2OH

2

rhamnose

2

HOOH

OH

OH

4OH

1308

21OHHOHHOH

HOH

HO

CH2OH

OHxylitolxylose

1
HOH2C

5HOOH4CH2OH

5OHO

333CH2OH

HOH

HOH

HOH

Hfructose

OH
CH2OH

CH2OH
OOH HO

4D-sorbitol
OH

OH

mannose
CH2OH
OOH

54510 min

OH

HOOH

myo-inositol

OH

HO0OH

HOHO0OH

glucose510Unison UK-Amino, 250 x 2 mm
acetonitrile /water = 80 /20, 0.2 mL/min
37 deg.C, ELSD

Unison UK-Amino, 250 x 2 mm
acetonitrile /water = 80 /20, 0.2 mL/min, 37 deg.C, ELSD

2CH2OHO
OH

15min
HOCH2OH
OH
OH
OHO
OH

5maltitol

OHNormal Phase Separation of Saccharides
Sorbitol and Mannitol
313

1 2

Unison UK-Amino, 250 x 2 mm
0.2 mL/min (5MPa), 0.5 uL (2.5ug)

Sharp peak shape is possible
with sugar alcohols by raising
the column temperature settings.

1CH2OH
HOH

HOH

H

OH

H

OH
CH2OH

1 2

D-sorbitol

Unison UK-Amino, 250 x 4.6 mm
1 mL/min (6MPa), 1.5 uL (7.5ug)
2CH2OH
HOH

HO

H

H

OH

H

OH
CH2OH

D-mannitol

21Brand-N (5um), 250 x 4.6mm
1 mL/min (4MPa), 1.5 uL (7.5ug)

010

acetonitrile / water = 90 / 1050 deg.C, ELSD

20 minArabinose, Galactose, Maltitol
320

Sucrose and Lactose in Cafe au Lait
321

311220510 min

0Unison UK-Amino, 250 x 4.6 mm
acetonitrile /water = 75 /25, 1 mL/min, 37 deg.C, ELSD

1O
HOHO

2OH

CH2OH
HOO
OH

OH

OH

D-galactose

CH2OH
O
OH
HO

10 minUnison UK-Amino, 250 x 4.6 mm
acetonitrile /water = 75 /25, 1 mL/min, 37 deg.C, ELSD

3OH

D-arabinose

51
CH2OH
OH
OH
OH

O
OH

CH2OH
O
OH
HOOH

OH

2CH2OHO
HOO

sucrosemaltitol

3CH2OH

HOCH2OHO OOHCH2OH
O
OH

OH
OH

OH
OH

lactoseNormal Phase Separation of Saccharides
Reducing Sugar and its Reduced Alcohl
315

112

CH2OH
H

OH

HO

H

H

OH

H

OH

3
CH2OH
O
OH

4
CH2OH
O
OH

OH

CH2OH
OH
OH

HO

HO

OH

O
OH

OH

CH2OH
O

CH2OH
O

OH

OH

HO

OH

O
OH

OH

OH

CH2OH

D-sorbitol

D-glucose

maltitolmaltoseUnison UK-Amino, 250 x 4.6 mm,
acetonitrile / water = 83 / 17, 1.0 mL/min, 50 deg.C, ELSD
23Cyclodextrins
310

405

1015 min

Unison UK-Amino, 250 x 2 mm
acetonitrile / water = 70 / 300.2 mL/min, 37 deg.C, ELSD

051015 min

Laminariorigosaccharides
Unison UK-Amino provides exceptional separation
efficiency for oligosaccharides and other chemicalcompounds with relatively higher molecular weights.

306

1

CH2OH
O
OH

1

OH

HOOH

glucose
2
3

6

4

CH2OH
O

7

OH
HO

5

O

CH2OH
O

HO
OH

O

CH2OH
O
OH

HOOH

nOH

2. laminaribiose (n=0)
3. laminaritriose (n=1)
4. laminaritetraose (n=2)
5. laminaripentaose (n=3)
6. laminarihexaose (n=4)
7. laminariheptaose (n=5)

0510Unison UK-Amino, 250 x 2 mm
A: acetonitrile, B: water, 25-40 %B (0-15min)
0.2 mL/min, 37 deg.C, ELSD15 min4

Aqueous Normal Phase Separation
Unison UK-Amino can conduct aqueous normal phase separatin even with chemicalcompounds other than carbohydrates. The column can optimally handle various compoundswith its combination of electrostatic interactions and anion exchange mode. In that case, LCUV/VIS, LC-ELSD and LC-MS is possible by optimizing the organic solvent strength and type,and by adjusting the buffer pH and ionic strength.
Branched-Chain Amino AcidsIn amino acid separation with both electrolytes, sharp peak shape ispossible by controlling pH and ion strength with neutral acetate
ammonium.

311

121H O

H2N

H3C
CH

CH2C3OHCH2CCOH

H3C

H3C

3H O

H2N

H3C

C

L-leucine

L-valine
H O

H2N

2

H3C

CH2CHC

COH

CH3

L-isoleucine0510 min

Unison UK-Amino, 150 x 2 mm
acetonitrile /10mM ammonium acetate = 85 /15
0.2 mL/min, 37 deg.C, ELSD

Ascorbic Acid and Erythorbic Acid
Ascorbic acid and its isomer erthyorbic acid can be separated in either normal phase or ion exchange modes. Unison
UK-Amino can be used with acetic acid, a mild pH adjusting agent. Moreover, separation mode differences allow
column users to select different elution orders and separation modes to suit their needs.
317

2 1

normal phase
Unison UK-Amino, 150 x 2 mm
acetonitrile /water /acetic acid = 80 /20 /2
0.2mL/min (5MPa),
37deg.C, 260nm,
1uL(1.3ug)

010

H
HOCH2OHCO

HOOH

Hanion exchange

HOCH2CHO

Unison UK-Amino, 250 x 2 mmwater / acetic acid = 100 / 0.5
0.2mL/min (14MPa),
37deg.C, 260nm,0.2uL(0.5ug)
10OH

1 L-ascorbic acid(Vitamin C)

20 min120O

20 min5OOOH

2 isoascorbic acid(erythorbic acid)Aqueous Normal Phase Separation (Water-soluble vitamins)
Water-Soluble Vitamins
309

Simple analysis is obtainable using acetic acid with watersoluble vitamins. There is no need for ion-pair mode via
reversed-phase separation. Moreover, gradient elution
enables high speed analysis for a wide range of vitamins

2O

1CNH2CH2N+1N

nicotinamide
4NH23N2510N

CH2CH2OH

Sthiamine

60H3C

CH320 min

Unison UK-Amino, 100 x 4.6 mm
ACN / water / acetic acid = 90 / 10 / 51mL/min, 37 deg.C, 260 nm

OH

OH

CH

CHCH2OHN

H3C

3OH
CH2CHNO
NH

N

H3C

O

riboflavinOC

4N

nicotinic acid

6OH

NH2COCH2CH2 CH3 CH3CH2CONH2
NH2COCH2
CH2CH2CONH2
H3C
N CNN
H3C
Co+2N

NH2COCH25N

CH3CH3H3C

HOO-

CH2CH2CONHCH2 C O P O

5cyanocobalamin

7CH3N

CH3CH3 CH2CH2CONH2H

1NHO

H3C

HHOHCH2OH

43L-ascorbic acid

05O

HOCH2OHCHOO

H6OH

10 minH2NUnison UK-Amin, 100 x 4.6 mm
A: ACN / acetic acid = 100 / 5
B: water / acetic acid = 100 / 52-70 %B (0-10min)
1mL/min, 37 deg.C, 260 nm

7NO

N

N
OH

folic acid

6N
CH2NH

O

C HNCH C
CH2CH2OHC OHO

Non-Aqueous Normal Phase Separation
Unison UK-Amino has a highly polar stationary phase enabling non-aqueous normal phaseseparation similar to silica columns. However, the presence of a dissociative group (aminogroup) and bound water in the stationary phase side means that highly reproducible analysis
is possible by adding acetic acid and other pH modifiers.
Steroids

Tocopherols
30731911218123495103476120510 min

0Unison UK-Amino, 250 x 4.6 mm
hexane / ethyl acetate / acetic acid = 80 / 20 / 0.1
1 mL/min, 37 deg.C, 295 nm

10Unison UK-Amino, 250 x 4.6 mm
A: hexane /acetic acid = 100 /0.1
B: ethanol /acetic acid = 100 /1
10-30%B (0-30 min), 1 mL/min, 37 deg.C, 260 nm

CH3OHO1OH3C30 min

20CH31CH2CH2CH2CH

CH3CH2OCCH3CH3CH32C OO

3C O

CH33HOO

O

progesterone

deoxycorticosterone
acetateestrone

CH3HOCH2OH

2OOH

CH3CH2CH2CH2CH

CH3CH3CH345 HOO

HOcorticosterone
CH2OH

H3C
CH3CH37

CH2CH2CH2CH
CH3C CH

HOestradiol

O

63HO3OHC OOCH3C O
OH

ethynylestradiol

CH2OH

8O

C O
OH

CH2OH

9HOC O
OH

3O

O

cortisoneO

6alphamethylprednisolone

prednisone

HO4OCH3CH3CH2CH2CH2CH
CH3CH3CH2OH

10HOC OOH

CH2OH11HOC OOH

12OH
OH

3O

HOO

hydrocortisone

7

prednisolone

estriolBatch-to-Batch Reproducibility
Unison UK-Amino stationary phase has an excellent batch-to-batch reproducibility fornormal phase + anion exchange mode.
1318

1

2

COOCH2CH2CH3

propylbenzoate

5OCH3HN

4O

2 HOCH23N

thymidine
pKa = 9.8O

HO34

COOH

benzoic acidpKa = 4.2

HNHO

NH2CO

3-hydroxyphenylurea

Conventional aminopropyl
stationary phases struggle
to achieve solute retention
and repeatable separations
as the interactions arecomplicated due to thepresence of both normal
phase and anion exchange
modes. Unison UK-Amino
addresses this problem witha novel stationary phasedesign to provides excellent
reproducibility.

O

UKAGC07DHNO

5HOCH2NOHOOH

uridinepKa = 9.2

UKAGD16E

UKAGD17E

05Unison UK-Amino, 150 x 2 mm
acetonitrile /water /acetic acid =95 /5 /1
0.2 mL/min (3.5MPa)37 deg.C, 270nm

10 minOrdering Information
Particle Size: 3um, Stationary Phase: Aminopropyl
Product Code / Price in JPY

Length(mm)1 mm I.D.

2 mm I.D.

3 mm I.D.

4.6 mm I.D.

6 mm I.D.

10 mm I.D.

1020305075
100150250500

UKA11 / 42,000
UKA12 / 45,000
UKA13 / 48,000
UKA14 / 53,000
UKA15 / 58,000
UKA16 / 75,000-

UKA20 / 45,000
UKA29 / 45,000
UKA21 / 38,000
UKA22 / 42,000
UKA23 / 45,000
UKA24 / 48,000
UKA25 / 53,000
UKA26 / 68,000-

UKA30 / 45,000
UKA39 / 45,000
UKA31 / 38,000
UKA32 / 42,000
UKA33 / 45,000
UKA34 / 48,000
UKA35 / 53,000
UKA36 / 68,000-

UKA00 / 45,000
UKA09 / 45,000
UKA01 / 38,000
UKA02 / 42,000
UKA03 / 45,000
UKA04 / 48,000
UKA05 / 53,000
UKA06 / 68,000
UKA07/110,000

UKA61 / 42,000
UKA62 / 45,000
UKA63 / 48,000
UKA64 / 53,000
UKA65 / 58,000
UKA66 / 75,000-

UKAP1 / 68,000
UKAP2 / 80,000
UKAP3 / 95,000
UKAP4 /120,000
UKAP5 /150,000
UKAP6 /180,000-

Guard Holder

Guard Cartridge UK-Amino
Sep. column I.D Prod.Code Price in JPY

Note

GCUKACGCUKASGCUKAM

5 x 1 mm, 3pcs
5 x 2 mm, 3pcs
10 x 8 mm, 2pcs

For 1 mm
For 2 - 6 mm
For 10 mm

23,00017,00023,000(With a column coupler)Sep. column I.D Prod.Code Price in JPY
GCH01S
26,000For 1 - 6 mm
For 10 mm

GCH02M

35,000

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