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Normal Phase Hydrophilic Interaction Chromatography

While reversed phase is the most widely used mode of retention in HPLC, very polar compounds are often not sufficiently retained in low percent organic, or even in 100% aqueous mobile phase. The order of elution in normal phase is opposite that found in RPC for the same mixture of compounds. Although non-polar organic mobile phases and a silica stationary phase were used traditionally in normal phase LC, today most normal phase separations are performed with aqueous-organic mobile phases and a more polar-bonded stationary phase. This mode of HPLC is now commonly referred to as HILIC, hydrophilic interaction chromatography.
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Description
Parameters

Normal Phase Hydrophilic Interaction Chromatography

While reversed phase is the most widely used mode of retention in HPLC, very polar compounds are often not sufficiently retained in low percent organic, or even in 100% aqueous mobile phase. The order of elution in normal phase is opposite that found in RPC for the same mixture of compounds. Although non-polar organic mobile phases and a silica stationary phase were used traditionally in normal phase LC, today most normal phase separations are performed with aqueous-organic mobile phases and a more polar-bonded stationary phase. This mode of HPLC is now commonly referred to as HILIC, hydrophilic interaction chromatography.

By using an amide or amino bonded phase column, polar compounds can be retained by a normal phase or hydrophilic interaction chromatography(HILIC; see references) retention mechanism using a mobile phase mixture of acetonitrile and ammonium acetate buffer. In contrast to the retention behavior in reversed phase, in HILIC, solutes will be retained longer when increasing the percent acetonitrile.

TSKgel Amide-80

TSKgel NH2-100

Bonded Phase

- Amide-80 bonded phase does
not react with reducing sugars.

- Anomer formation can be
prevented by raising mobile
phase temperature

up to 80°C.

- NH2-100 bonded phase is more stable than conventional amino phases due to a special endcapping prior to introduction of aminoalkyl groups.

- Amino-bonded phase can react with a reducing sugar to form a Schiff base.

Detectors

Can be used with evaporative light scattering (ELS) and mass spec(MS) detectors

Mobile Phase

Stable in 100% organic eluents

Solutes

Retains very polar drug candidates and drug metabolites

Applications

- saccharides

- polyols(polyalchols)

- drug metabolites

- peptides

- melanine & cyanuric acid

- oligonucleotides

- polar compounds in
- fermentation broth

cystosine

- saccharides

- polyols(polyalchols)

- methotrexate polyglutamate derivatives

- pyridylaminated oligosaccharides

- water-soluble vitamins

- nucleic acid fragments

TSKgel Amide-80 2µm

Particle Size(mean):

2 µm

Pore Size(mean):

10 nm

Functional Group:

carbamoyl

pH Range:

2.0 - 7.5

Max. temperature :

50 ºC

Surface Area(m2/g):

450

P/N

Description

Particle size

Housing material

ID(mm)

Length(cm)

0023454

TSKgel Amide-80 2µm

2 µm

Stainless Steel

2

5

0023455

TSKgel Amide-80 2µm

2 µm

Stainless Steel

2

10

0023456

TSKgel Amide-80 2µm

2 µm

Stainless Steel

2

15

0023457

TSKgel Amide-80 2µm

2 µm

Stainless Steel

3

5

0023458

TSKgel Amide-80 2µm

2 µm

Stainless Steel

3

10

0023459

TSKgel Amide-80 2µm

2 µm

Stainless Steel

3

15

0023460

TSKgel guardclumn Amide-80 2μm

2 µm

Stainless Steel

2

1

TSKgel Amide-80 3µm

Particle Size(mean):

3 µm

Pore Size(mean):

10 nm

Functional Group:

carbamoyl

pH Range:

2.0 - 7.5

Max. temperature :

50 ºC

Surface Area(m2/g):

450

P/N

Description

Particle size

Housing material

ID(mm)

Length(cm)

0021864

TSKgel Amide-80 3µm

3 µm

Stainless Steel

2

5

0021865

TSKgel Amide-80 3µm

3 µm

Stainless Steel

2

15

0022850

TSKgel Amide-80 3µm

3 µm

Stainless Steel

3

5

0022851

TSKgel Amide-80 3µm

3 µm

Stainless Steel

3

10

0022852

TSKgel Amide-80 3µm

3 µm

Stainless Steel

3

15

0021862

Guard Cartridge for 2 and 3mmID columns, 3pk

3 µm

Stainless Steel

2

1

0019308

Guard Cartridge Holder for P/N 0021862

Stainless Steel

2

1

0021866

TSKgel Amide-80 3µm

3 µm

Stainless Steel

4.6

5

0022849

TSKgel Amide-80 3µm

3 µm

Stainless Steel

4.6

10

0021867

TSKgel Amide-80 3µm

3 µm

Stainless Steel

4.6

15

0021863

Guard Cartridge for 4.6mmID columns, 3pk

3 µm

Stainless Steel

3.2

1.5

0019018

Guard Cartridge Holder for P/N 0021863

Stainless Steel

3.2

1.5

TSKgel Amide-80 5µm

Particle Size(mean):

5 and 10 µm

Pore Size(mean):

10 nm

Functional Group:

carbamoyl

pH Range:

2.0 - 7.5

Max. temperature :

80 ºC

Surface Area(m2/g):

450

P/N

Description

Particle size

Housing material

ID(mm)

Length(cm)

0020009

TSKgel Amide-80

5 µm

Stainless Steel

1

5

0020010

TSKgel Amide-80

5 µm

Stainless Steel

1

10

0021486

TSKgel Amide-80

5 µm

Stainless Steel

1

15

0021487

TSKgel Amide-80

5 µm

Stainless Steel

1

25

0019694

TSKgel Amide-80

5 µm

Stainless Steel

2

5

0019695

TSKgel Amide-80

5 µm

Stainless Steel

2

10

0019696

TSKgel Amide-80

5 µm

Stainless Steel

2

15

0019697

TSKgel Amide-80

5 µm

Stainless Steel

2

25

0021941

Guard Cartridge for 2mm ID columns, 3 pk, Replaces P/N 0042140

5 µm

Stainless Steel

2

1

0019308

Guard Cartridge holder for P/N 0021941

Stainless Steel

2

1

0019532

TSKgel Amide-80

5 µm

Stainless Steel

4.6

5

0019533

TSKgel Amide-80

5 µm

Stainless Steel

4.6

10

0013071

TSKgel Amide-80

5 µm

Stainless Steel

4.6

25

0019021

Guard Column for 4.6mm ID columns

5 µm

Stainless Steel

4.6

1

0019010

Guard Column for 4.6mm ID columns, 3 pk

5 µm

Stainless Steel

3.2

1.5

0019018

Guard Cartridge holder for P/N 0019010

Stainless Steel

3.2

1.5

0014459

TSKgel Amide-80

10 µm

Stainless Steel

7.8

30

0014460

TSKgel Amide-80

10 µm

Stainless Steel

21.5

30

0014461

Guard column for P/N 0014460

10 µm

Stainless Steel

21.5

7.5

TSKgel NH2-100 3µm

Particle Size(mean):

3 µm

Pore Size(mean):

10 nm

Functional Group:

alkylamine

pH Range:

2.0 - 7.5

Max. temperature:

50 ºC

Surface Area(m2/g):

450

P/N

Description

Particle size

Housing material

ID(mm)

Length(cm)

0021967

TSKgel NH2-100

3 µm

Stainless Steel

2

5

0021968

TSKgel NH2-100

3 µm

Stainless Steel

2

15

0021971

Guard Cartridge for 2mmID columns, 3pk

3 µm

Stainless Steel

2

1

0019308

Guard Cartridge Holder for P/N 21971

Stainless Steel

2

1

0021999

TSKgel NH2-100 DC

3 µm

Stainless Steel

4.6

5

0021969

TSKgel NH2-100

3 µm

Stainless Steel

4.6

5

0021970

TSKgel NH2-100

3 µm

Stainless Steel

4.6

15

0021972

Guard Cartridge for 4.6mmID columns, 3pk

3 µm

Stainless Steel

3.2

1.5

0019018

Guard Cartridge Holder for P/N 21972

Stainless Steel

3.2

1.5
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