columns and bulk packings for bio-macro molecular ... general catalog 5 columns and bulk packings...

YMC GENERAL CATALOG

5Columns and bulk packingsfor BIO-MACROMolecular Separations

Types and characteristics of columns and bulk packing for BIO-MACRO molecular separations

YMC-BioPro Ion Exchange columnsYMC-BioPro QA-F／YMC-BioPro SP-FYMC-BioPro QA／YMC-BioPro SPYMC-BioPro Ion Exchange mediaYMC-Pack DiolReversed-phase columns for bio moleculesYMC-Pack PROTEIN-RPYMC-Pack Diol-NPYMC-Pack Polyamine Ⅱ

102～105

106107

108,109110,111112,113

114～121122,123

124125

101

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olecular Separations

Columns and bulk packings for BIO-MACRO molecular separationsIt is increasingly more important to separate bio molecules such as peptides, proteins and nucleic acids, due to progress of bio pharmaceuticals like monoclonal antibodies. To meet such demands, YMC offers abundant columns and bulk packings for ion exchange, size exclusion, reversed-phase and normal-phase chromatography.Depending on target compounds or scale of the separation, choose appropriate columns and packings.

Columns and bulk packings for BIO-MACRO separations　(See P10, 11 for choosing appropriate column for target compounds.)

Separation mode Product name Base

materialPore size

（Å） Functional group Purpose compounds Page

Ion exchange

YMC-BioPro QA-F

Polymer

non-porous−CH2N＋（CH3）3

ProteinsPeptidesNucleic acidsOligonucleotides

106 〜111

YMC-BioPro SP-F −CH2CH2CH2SO3−

YMC-BioPro QAporous

−CH2N＋（CH3）3

YMC-BioPro SP −CH2CH2CH2SO3−

YMC-BioPro Q30YMC-BioPro Q75

porous−CH2N＋（CH3）3

YMC-BioPro S30YMC-BioPro S75 −CH2CH2CH2CH2SO3

−

Size exclusion YMC-Pack Diol Silica gel 60,120, 200, 300 Dihydroxypropyl

ProteinsPeptidesNucleic acidsOligonucleotidesCarbohydrates

112,113

Reversed-phase

Triart C18 Hybrid silica 120

C18ProteinsPeptidesNucleic acidsOligonucleotidesNucleotidesNucleosidesNucleobasesCarbohydrates

114 〜121

YMC-Pack Pro C18

Silica gel

120Hydrosphere C18 120YMC-Pack Pro C18 RS 80YMC-Pack ODS-A 120, 200, 300YMC-Pack ODS-AQ 120, 200YMC-Pack C8 120, 200, 300 C8YMC-Pack C4 120, 200, 300 C4YMC-Pack Ph 120 PhenylYMC-Pack CN 120, 300 CyanopropylYMCbasic 200 C8YMC-Pack PROTEIN-RP 200 − 122,123YMC-Pack PolymerC18 Polymer − C18 114 〜121

Normal phase

YMC-Pack Diol-NP

Silica gel

60,120 Dihydroxypropyl PeptidesNucleotidesNucleosidesNucleobasesCarbohydrates

124

YMC-Pack Polyamine Ⅱ 120 Polyamine 125

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0 10 20 30 min

Transferrin

Albumin

N080313E

IgGmAU

10

0

YMC-BioPro QA 5 µm, 50×4.6 mmI.D.

0 10 20 30 40 50 60 min

N080616A

IgG

AlbuminTransferrin

mAU

150

100

0

50

YMC-Pack Diol-300 + Diol-200 5 µm,　300×8.0 mmI.D. ×2

Eluent ： A) 20 mM Tris-HCl (pH 8.6) B) 20 mM Tris-HCl (pH 8.6) containing 0.5 M NaCl 0-30% B (0-15 min), 30-100% B (15-30 min)

Flow rate ：0.5 mL/minTemperature ：25 ℃Detection ：UV at 280 nmInjection ：20 µL (100 µL/mL)

Eluent ：0.1 M KH2PO4-K2HPO4 (pH 7.0) containing 0.2 M NaClFlow rate ：0.5 mL/minTemperature ：ambient (25 ℃)Detection ：UV at 280 nmInjection ：20 µL (100 µL/mL)

Proteins in human serum are separated by the difference in the surface charge on ion exchange chromatography (IEC) and by the difference in the molecular weight on size exclusion chromatography (SEC).

Separation of proteins Comparison of separation in different modeHuman serum

Ion exchange Size exclusion

min2 4 6 8 10 12 14 160

NaN3

Mouse monoclonal IgG1(Anti-human IgG4)

mAU

0

1

2

3

4

5

P080220A

YMC-BioPro QA-F 5 µm, 30×4.6 mmI.D.

0 105 15 20 25 30 min

NaN3Mouse monoclonal IgG1(Anti-human IgG4)

0

10

20

30

40

mAU

P080530A

YMC-Pack Diol-200 5 µm, 300×4.6 mmI.D.

Eluent ： A) 20 mM Tris-HCl (pH 8.1) B) 20 mM Tris-HCl (pH 8.1) containing 0.5 M NaCl 10-25% B (0-18 min)

Flow rate ：1.0 mL/minTemperature ：25 ℃Detection ：UV at 220 nmInjection ：10 µL (0.1 mg/mL)

Eluent ：0.1 M KH2PO4-K2HPO4 (pH 7.0)Flow rate ：0.17 mL/minTemperature ：ambient (25 ℃)Detection ：UV at 220 nmInjection ：10 µL (0.05 mg/mL)

Mouse monoclonal antibody against human IgG4 is analyzed on ion exchange chromatography (IEC) and size exclusion chromatography (SEC). Several peaks possibly derived from isoform of antibody are observed in ion exchange mode, while a single peak is detected in size exclusion mode.

Mouse monoclonal IgG1 anti-human IgG4 (Purified by DEAE chromatography, containing NaN3)

1. Rabbit IgG2. Mouse IgG Fc fragment

min

mAU

0 5 10 15 20

0

25

50

R080623D

1

2


30 40352520151050 min

R080619B

Mouse IgG Fc fragmentmAU

0

10

20

30

YMC-Pack C4（300 Å）5 µm, 150×4.6 mmI.D.

Eluent ：0.1 M KH2PO4-K2HPO4 (pH 6.9) containing 0.2 M NaClFlow rate ：1.0 mL/minTemperature ：ambient (27 ℃)Detection ：UV at 220 nmInjection ：5 µL (0.5 mg/mL)

Eluent ： A) water/TFA (100/0.1) B) acetonitrile/TFA (100/0.1) 25-45% B (0-40 min)

Flow rate ：1.0 mL/minTemperature ：37 ℃Detection ：UV at 220 nmInjection ：5 µL (1.0 mg/mL)

Size exclusion chromatography (SEC) is useful for separation of substances which have distinct differences in molecular weight, like between IgG and its fragments. On the other hand, reversed-phase chromatography (RPC) is suitable for a precise analysis of peptides and proteins with a molecular weight of less than 100 kDa such as IgG Fc fragment.

Mouse IgG Fc fragment (Prepared from normal serum)

Ion exchange Size exclusion

Size exclusion Reversed-phase

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Columns and bulk packings for BIO-MACRO molecular separations

0 60 min2010 30 40 50

N080422B

*

0

10

20

mAU

YMC-BioPro QA 5 µm, 50×4.6 mmI.D.


Flow rate ：0.5 mL/minTemperature ：25 ℃Detection ：UV at 220 nmInjection ：20 µL

Separation of proteins Comparison of separation in different modeTryptic digests of BSA

Ion exchange

0 60 min2010 30 40 50

0

30

50

mAU

60

40

20

10

N080623L

105

102

103

104

MW

5

4

3

2

1

101*

YMC-Pack Diol-120 + Diol-60 5 µm, 500×8.0 mmI.D. ×2

These chromatograms show separation of tryptic digests of BSA (MW: 66,000) in ion exchange chromatography (IEC), size exclusion chromatography (SEC) and reversed-phase chromatography (RPC). The molecular weight of the digests is estimated to be approximately from 100 to 20,000 by SEC chromatogram. IEC and RPC chromatograms show many peaks of fragments which are separated by the difference in structure, charge and hydrophobicity.

Size exclusion

Eluent ： 0.1 M KH2PO4-K2HPO4 (pH 7.0) containing 0.2 M NaCl/acetonitrile (70/30)

Flow rate ：0.7 mL/minTemperature ：ambient (25 ℃)Detection ：UV at 220 nmInjection ：5 µL

0 60 min2010 30 40 50

0

20

30

mAU

10

40

N080318B

*

* undigested BSA

YMCbasic 5 µm, 150×2.0 mmI.D.Reversed-phase

Eluent ： A) water/TFA (100/0.1) B) acetonitrile/TFA (100/0.1) 5-35% B (0-50 min), 35-45% B (50-55 min), 45% B (55-60 min)


Calibration curve of peptides and proteins1. Myoglobin (MW 17,000)2. Insulin (Bovine) (MW 5,700)3. Neurotensin (MW 1,672)4. Tetraglycine (MW 246)5. Glycine (MW 75)

20 min100G910607A

YMC-Pack ODS-A（120 Å）5 µm,150×4.6 mmI.D.

Separation of sugar chains Comparison of separation in different modePyridylamino (PA) -Sugar chains

Reversed-phase

20 min100G910620A

YMC-Pack Polyamine Ⅱ 5 µm,150×4.6 mmI.D.

Pyridylamino (PA) sugar chains are often analyzed for structural determination of sugar chain in glycoproteins and glycolipids. Separations of PA sugar chains in reversed-phase (RP) mode and normal-phase (NP) mode are shown. Two dimensional HPLC combining two different modes, such as RP mode and NP mode, is useful tool for structural determination of sugar chain.

Normal-phase

Eluent ：methanol/20 mM NH4H2PO4 (5/95)Flow rate ：1.0 mL/minTemperature ：37 ℃Detection ：FLS at Ex. 320 nm, Em. 400 nmInjection ：2 µL (3.3 pmol/mL)Sample ： PA-Sugar Chain Series,

manufactured by TAKARA SHUZO CO., LTD.

Eluent ：methanol/20 mM NH4H2PO4 (80/20)Flow rate ：1.0 mL/minTemperature ：37 ℃Detection ：FLS at Ex. 320 nm, Em. 400 nmInjection ：3 µL (3.3 pmol/mL)Sample ： PA-Sugar Chain Series,

manufactured by TAKARA SHUZO CO., LTD.

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N080212G

0 2010 30 min

0

50

100

mAU


Separation of nucleic acids Comparison of separation in different modeDNA fragments 1 Kb DNA ladder (75 - 12,216 bp)

Ion exchange

Eluent ： A) 20 mM Tris-HCl (pH 8.1) containing 0.7 M NaCl B) 20 mM Tris-HCl (pH 8.1) containing 1.0 M NaCl 0-100% B (0-30 min)


DNA fragments are analyzed with YMC-BioPro QA-F ion exchange column. 100 mm length column of YMC-BioPro QA-F is ideal for high-resolution analysis of nucleic acids.


0 10 20 25 min155

N080610F

Plasmid pBR322 Hae III digests (8-587 bp)

Plasmid pBR322 (4,361 bp)

10

20

30

40

mAU

0

Plasmid pBR322 restriction fragments

Ion exchange

Eluent ： A) 20 mM Tris-HCl (pH 8.1) B) 20 mM Tris-HCl (pH 8.1) containing 1.0 M NaCl 70-85% B (0-20 min), 85% B (20-25 min)


YMC-Pack Diol-300 + Diol-200　5 µm, 500×8.0 mmI.D. ×2

P080617A

Plasmid pBR322 Hae III digests (8-587 bp)

Plasmid pBR322 (4,361 bp)

10

20

30

40

mAU

0

0 20 403010 50 min

Size exclusion

Eluent ：0.1 M KH2PO4-K2HPO4 (pH 7.0) containing 0.2 M NaClFlow rate ：0.7 mL/minTemperature ：ambient (25 ℃)Detection ：UV at 260 nmInjection ：10 µL

The separation of plasmid pBR322 restriction fragments (8-857 bp) is compared between in ion exchange mode and size exclusion mode. Ion exchange chromatography (IEC) is applicable to identification of each fragment requiring high resolution and size exclusion chromatography (SEC) is usable for characterization of molecular weight distribution.

5'-CCGCTCGAGCTAAAAAAAGCCTGTGTTACC-3' (30 mer)

0 5 10 15 20 min

UV

TIC

30 mer2928

27

F060213C-04

Hydrosphere C18 3 µm, 50×2.0 mmI.D.

Primer of DNA sequencing (30 mer)

Reversed-phaseEluent ： A) 10 mM DBAA* (pH 6.0)

B) 10 mM DBAA* (pH 6.0)/acetonitrile (50/50) 58-62% B (0-20 min), 62% B (20-25 min)

Flow rate ：0.2 mL/minTemperature ：35 ℃Detection ：UV at 269 nm and ESI negative-modeInjection ：1 µL (10 pmol/component)* di-n-butylammonium acetate

This figure shows LC/MS analysis of a mixture of synthetic 27-30 mer oligonucleotides in reversed-phase mode. Hydrosphere C18 columns have been designed for enhanced retention and selectivity of highly polar compounds. They can achieve excellent separation by one-nucleotide difference and sufficient intensity in UV and ESI-MS.

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Ion Exchange Columns

YMC-BioPro Ion Exchange ColumnsYMC-BioPro ion exchange columns are specially designed for separation of proteins, peptides, and nucleic acids. YMC-BioPro ion exchange columns are available in QA and SP chemistries and are based on 5 µm porous and non-porous hydrophilic polymer beads with low nonspecific adsorption.

Ion exchange columns ideal for analysis of proteins, peptides, and nucleic acids

Features

● Ion exchange columns designed for analytical and laboratory-scale purification of proteins, peptides, and nucleic acids

● Newly developed hydrophilic polymer beads with low nonspecific adsorption

● Effective surface structure designed for maximum interaction with biomolecules

● Available in a strong anion exchanger (QA, quaternary ammonium) and a strong cation exchanger (SP, sulfopropyl)

● Non-porous type for increasing resolution and throughput

● Porous type for higher binding capacity and recovery

SpecificationsYMC-BioPro QA-F YMC-BioPro SP-F YMC-BioPro QA YMC-BioPro SP

Matrix Hydrophilic non-porous polymer Hydrophilic porous polymerParticle size 5 µmCharged group −CH2N＋（CH3）3 −CH2CH2CH2SO3

− −CH2N＋（CH3）3 −CH2CH2CH2SO3−

Counter ion Cl− Na＋ Cl− Na＋

Ion exchange capacity（meq/mL-resin）

0.075 - 0.110 0.230 - 0.290 0.075 - 0.100 0.070 - 0.095

Dynamic binding capacity（mg/mL-resin）

>12（BSA） >10（human-IgG） >110（BSA） >70（human-IgG）

Usable temperature 4 - 60 ℃Usable pH range 2.0 - 12.0Column material PEEK

SEM images of polymer beads of YMC-BioPro ion exchange columns

Porous polymer beadsNon-porous polymer beads

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YMC-BioPro QA-F ／ YMC-BioPro SP-F● Ion exchange column based on non-porous polymer beads● High efficiency with low operating pressure● 30 mm length column for ultra high-throughput analysis● 100 mm length column for high-resolution analysis

Ion exchange columns for high-throughput and high-resolution analysis of proteins, peptides, and nucleic acidsYMC-BioPro QA-F/SP-F columns are ion exchange columns based on non-porous hydrophilic polymer beads with high chemical and mechanical stability, and low nonspecific adsorption of biomolecules. The short

columns (30 mm, 50 mm) are useful for the fast analysis at a higher flow rate, and the 100 mm length columns are best choice for the quality control assessment of biopharmaceuticals requiring a high-resolution.

■Matrix：Hydrophilic non-porous polymer beads■Usable pH range：2.0 〜 12.0

Ultra high-throughput analysis of proteins

YMC-BioPro SP-F 5 µm, 30×4.6 mmI.D.YMC-BioPro SP-F 5 µm, 30×4.6 mmI.D.

1 2

3

N071023K

1 2 3 4 min0

0.0

0.2

0.4

AU1. Ribonuclease A2. Cytochrome c 3. Lysozyme

The high mechanical stability of non-porous polymer beads and the short column length enable faster elution of proteins at a higher flow rate.

Eluent ： A) 20 mM KH2PO4-K2HPO4 (pH 6.8) B) 20 mM KH2PO4-K2HPO4 (pH 6.8) 　 containing 0.5 M NaCl 0-100% B (0-4 min)

Flow rate ：1.5 mL/min (540 cm/hr)Temperature ：25 ℃Detection ：UV at 220 nmInjection ：20 µLPressure ：4.8-5.2 MPa

Packing material

Particle size（µm）

Pore size（Å）

Column sizeinner diameter×length（mm） Product number

QA-F S-5 non-porous

4.6× 30 QF00S05-0346WP4.6× 50 QF00S05-0546WP4.6×100 QF00S05-1046WP

Packing material


Pore size（Å）


SP-F S-5 non-porous

4.6× 30 SF00S05-0346WP4.6× 50 SF00S05-0546WP4.6×100 SF00S05-1046WP

Ordering information

High-resolution analysis of nucleic acids

YMC-BioPro QA-F 5 µm, 100×4.6 mmI.D.YMC-BioPro QA-F 5 µm, 100×4.6 mmI.D.

N080212G

20 min100

50

mAU

0The separation of DNA fragments is shown. YMC-BioPro QA-F of 100 mm length column is good choice for high-resolution analysis of nucleic acids.

Eluent ： A) 20 mM Tris-HCl (pH 8.1) containing 0.7 M NaCl B) 20 mM Tris-HCl (pH 8.1) containing 1.0 M NaCl 0-100% B (0-30 min)

Flow rate ：0.5 mL/min (180 cm/hr)Temperature ：25 ℃Detection ：UV at 260 nmInjection ：20 µL (0.25 mg/mL)

DNA fragments 1Kb DNA ladder (75 - 12,216 bp)

High-resolution analysis of proteins

YMC-BioPro QA-F 5 µm, 100×4.6 mmI.D.YMC-BioPro QA-F 5 µm, 100×4.6 mmI.D.

P080424A

2010 30 40 min5 15 25 35

P080424B

20100 30 40 min5 15 25 35

NaN3

NaN3

MAb Lot. A

MAb Lot. B MAb

MAb

mAU

0

1

2

3

4

mAU

0

1

2

3

4 Two different lots of commercially available MAb purified by DEAE chromatography, are analyzed with 100 mm length column of YMC-BioPro QA-F. The MAb is resolved into several peaks, and the lot-to-lot variability is observed. 100 mm length column of YMC-BioPro QA-F/SP-F, which has high efficiency, is ideal for characterization of glycoproteins such as monoclonal antibodies and for quality control assessment of biopharmaceuticals.

Eluent ： A) 20 mM Tris-HCl (pH 8.1) B) 20 mM Tris-HCl (pH 8.1) containing 0.5 M NaCl 10-25% B (0-60 min)

Flow rate ：1.0 mL/min (360 cm/hr)Temperature ：25 ℃Detection ：UV at 220 nmInjection ：14 µL (0.1 mg/mL)Sample ： Mouse monoclonal IgG1 anti-human IgG4

(Purified by DEAE chromatography, containing NaN3)

Monoclonal antibody (MAb) against human IgG4

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YMC-BioPro QA ／ YMC-BioPro SP● Ion exchange column based on porous polymer beads● Excellent resolution● High binding capacity and high recovery of biomolecules● Suitable for laboratory-scale purification

Ion exchange columns for analysis and laboratory-scale purification of proteins, peptides, and nucleic acidsYMC-BioPro QA/SP columns are ion exchange columns based on porous hydrophilic polymer beads with low nonspecific adsorption of biomolecules. YMC-BioPro QA/SP columns have superior resolution, high binding

capacity and high recovery of various biomolecules, and they allow highly effective analysis and laboratory-scale purification of biopharmaceutical proteins such as antibodies.

■Matrix：Hydrophilic porous polymer beads■Usable pH range：2.0 〜 12.0

Excellent resolutions

The separation of standard protein mixture is compared between YMC-BioPro SP and a commercial porous polymer cation exchange column. Many impurities are resolved from the main peaks of proteins on YMC-BioPro SP with superior peak shapes.

Eluent ： A) 20 mM KH2PO4-K2HPO4 (pH 6.8) B) 20 mM KH2PO4-K2HPO4 (pH 6.8) 　 containing 0.5 M NaCl 0-100% B (0-60 min)

Flow rate ： 0.5 mL/min (180 cm/hr)Temperature ：25 ℃Detection ：UV at 220 nmInjection ： 20 µL

1. Ribonuclease A (0.5 mg/mL)2. Cytochrome c (0.5 mg/mL)3. Lysozyme (0.5 mg/mL)

0 20 40 60 min

0

0.5

AU

1 23

YMC-BioPro SP5 µm, 50×4.6 mmI.D.

1 2 3

N071016E

0 20 40 60 min

0

0.5

AU

1 23

Brand T (porous S type)7 µm, 50×4.6 mmI.D.

1 2 3

N071018F

0 10 20 30 min

mAU

10

0

IgG

Albumin

N080303G

Transferrin

0 10 20 30 min

mAU

10

0

Transferrin

Albumin

N080313E

IgG

0 10 20 30 min

mAU

10

0 N080304A

Transferrin

AlbuminIgG

YMC-BioPro QA5 µm, 50×4.6 mmI.D.

Brand G (porous Q type)10 µm, 50×5.0 mmI.D.

Brand T (porous Q type)10 µm, 50×4.6 mmI.D.


Flow rate ：0.5 mL/min (180 cm/hr for 4.6 mmI.D., 150 cm/hr for 5.0 mmI.D.)Temperature ：25 ℃Detection ：UV at 280 nmInjection ：20 µLSample ：Human serum (100 µL/mL)

Separation of proteins in human serumComparison of separation on YMC-BioPro QA and commercial porous Q type columns

YMC-BioPro QA5 µm, 50×4.6 mmI.D.

Brand G (porous Q type)10 µm, 50×5.0 mmI.D.

Brand T (porous Q type)10 µm, 50×4.6 mmI.D.

0 2010 30 40 50 60 min

0 2010 30 40 50 60 min

0 2010 30 40 50 60 min

peaks = 50

peaks = 11

peaks = 28

mAU

30

20

0

10

mAU

30

20

0

10

mAU

30

20

0

10

N080422B

N080417G

N080417C


Flow rate ：0.5 mL/min (180 cm/hr for 4.6 mmI.D., 150 cm/hr for 5.0 mmI.D.)Temperature ：25 ℃Detection ：UV at 220 nmInjection ：20 µLSample ：Tryptic digest of BSA

Peptide mappingComparison of separation on YMC-BioPro QA and commercial porous Q type columns

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Analytical columnsPacking material


Pore size（Å）


QA S-5porous

4.6× 30 QAA0S05-0346WP4.6× 50 QAA0S05-0546WP4.6×100 QAA0S05-1046WP

Packing material


Pore size（Å）


SP S-5porous

4.6× 30 SPA0S05-0346WP4.6× 50 SPA0S05-0546WP4.6×100 SPA0S05-1046WP


High loadabilityComparison of the effect of sample load on YMC-BioPro QA and commercial Q type column

1

2

N080206A

N080206B

N080207C

N080207B

500 µg

250 µg

100 µg

50 µg

Loading amount

20100 30 min

12

N080206D

N080206E

N080208C

N080208B

20100 30 min

500 µg

250 µg

100 µg

50 µg0

100

200

mAU

0

100

200

mAU

YMC-BioPro QA5 µm, 50×4.6 mmI.D.YMC-BioPro QA5 µm, 50×4.6 mmI.D.

Brand G (porous Q type)10 µm, 50×5.0 mmI.D.Brand G (porous Q type)10 µm, 50×5.0 mmI.D.

YMC-BioPro QA shows the excellent resolution and peak shapes even when the loading amount increases. The porous type YMC-BioPro columns are suitable for laboratory-scale purification of proteins.

Eluent ： A) 20 mM Tris-HCl (pH 8.1) B) 20 mM Tris-HCl (pH 8.1) 　 containing 0.5 M NaCl 10-80% B (0-30 min)

Flow rate ： 0.5 mL/min (180 cm/hr for 4.6 mmI.D., 150 cm/hr for 5.0 mmI.D.)

Temperature ：25 ℃Detection ：UV at 280 nmInjection ：100 µL

1. Ovalbumin2. Trypsin inhibitor

High binding capacity and recoveryComparison of dynamic binding capacity (DBC) and recovery for BSA

Dynamic binding capacity（mg/mL-resin, 10% breakthrough）

Eluted amount（mg/mL-resin）

Recovery*（%）

YMC-BioPro QA 126 120 95Brand T (porous Q type) 73 58 79Brand G (porous Q type) 100 35 35

*Recovery：（Eluted amount/Dynamic binding capacity）×100

YMC-BioPro QA gives the superior DBC and recovery compared with conventional porous polymer anion exchange columns. The surface structure of YMC-BioPro which is designed for maximum interaction with proteins provides high binding capacity, and the hydrophilic property of polymer beads remarkably reduces nonspecific adsorption of proteins

Column ： YMC-BioPro QA 50×4.6 mmI.D. Brand T (porous Q type) 50×4.6 mmI.D. Brand G (porous Q type) 50×5.0 mmI.D.

Linear velocity ：180 cm/hrEquilibration buffer ：20 mM Tris-HCl (pH 8.6)Elution buffer ：20 mM Tris-HCl (pH 8.6) containing 1.0 M NaClSample ：1 mg/mL Bovine serum albumin (BSA) in equilibration bufferDetection ：UV at 280 nm

Breakthrough curves

10% breakthrough

BSA amount loaded (mg/mL-resin)

0 20 40 60 80 100 120 140 160

Brand T

Brand G

BioPro QA

0

0.50

0.25

AU

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Ion Exchange Media

YMC-BioPro Ion Exchange Media● Hydrophilic polymer beads with low nonspecific adsorption● Strong anion exchanger (Q type) and strong cation exchanger (S type)● Effective surface structure designed for maximum interaction with biomolecules● High binding capacity and recovery

Ion exchange media for capture and intermediate purification of biomoleculesYMC-BioPro ion exchange media are available in Q and S chemistries on 75 µm and 30 µm of hydrophilic porous polymer beads with low nonspecific adsorption and high binding capacity. YMC-BioPro ion exchange

media have similar retention selectivity to YMC-BioPro QA/SP columns, and it allows predictable scale up from analytical to preparative separation.

■Matrix：Hydrophilic porous polymer beads■Usable pH range：2.0 〜 12.0

SpecificationsYMC-BioPro Q30 YMC-BioPro Q75 YMC-BioPro S30 YMC-BioPro S75

Matrix Hydrophilic porous polymer beadsParticle size 30 µm 75 µm 30 µm 75 µmCharged group −CH2N＋（CH3）3 −CH2CH2CH2CH2SO3

−

Usable pH range 2.0 - 12.0

High dynamic binding capacity (DBC) for proteins

Anion exchanger Particle size（µm）


DBC*1

（mg/mL-resin）YMC-BioPro Q75 75 0.13 183Brand G（porous Q type） 90 0.19 102

Cation exchanger Particle size（µm）


DBC*1

（mg/mL-resin）YMC-BioPro S75 75 0.12 192Brand G（porous S type） 90 0.13 80

YMC-BioPro ion exchange media have 1.4 to 1.8 times higher DBC of protein than commercial ion exchange media. YMC-BioPro ion exchange media are effective in protein purification from capture step requiring high capacity to intermediate step requiring high efficiency.

*1 Dynamic binding capacities were determined at 10% breakthrough under following conditions:

Column ： 50×4.6 mmI.D.Linear velocity ：180 cm/hr

for anion-exchange resinEquilibration buffer ：20 mM Tris-HCl (pH 8.6)Elution buffer ：0.5 M NaCl in equilibration bufferSample ：1.5 mg/mL BSA in equilibration bufferDetection ：UV at 280 nm

for cation-exchange resinEquilibration buffer ：20 mM Glycine-NaOH (pH 9.0)Elution buffer ：0.5 M NaCl in equilibration bufferSample ：1.5 mg/mL Lysozyme in equilibration bufferDetection ：UV at 300 nm

160

180

200

220

240

260

0 200 400 600 800 1000 1200

YMC-BioPro S75Brand T（porous S type）Brand G（porous S type）

DBC（mg/mL-resin）

Linear velocity（cm/hr）

Dependency of DBC to linear velocity

YMC-BioPro ion exchange media show high DBC over a wide range of linear velocity, and the difference of DBC is less than 5% between 200 cm/hr and 1000 cm/hr. YMC-BioPro ion exchange media give increased productivity and reduced cost in biopharmaceutical production.

Column ：50×5.0 mmI.D.Equilibration buffer ：20 mM Glycine-NaOH (pH 9.0)Elution buffer ：0.5 M NaCl in equilibration bufferSample ：1.0 mg/mL Lysozyme in equilibration bufferDetection ：UV at 300 nm

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Excellent durability under CIP condition with 1 M NaOH

Cleaning in place (CIP) is an important procedure for cleaning and sterilization of columns used for protein purification. The DBC and the selectivity of proteins are unaffected following 20 cycles of CIP with 1 M NaOH. The high chemical stability of BioPro ion exchange media allows effective cleaning with alkaline solution.

Conditions of DBC measurementColumn ：YMC-BioPro S75 50×5.0 mmI.D.Linear velocity ：800 cm/hrEquilibration buffer ：20 mM Glycine-NaOH (pH 9.0)Elution buffer ：0.5 M NaCl in equilibration bufferSample ：1.0 mg/mL Lysozyme in equilibration bufferDetection ：UV at 300 nm※DBC is determined at 10% breakthrough

Test protocols

Separation of standard proteins

Determination of DBC and recovery

CIP with 1 M NaOH(5 column volumes at 400 cm/hr)

DBC and recovery

Number of CIP with 1M NaOH

120

100

60

40

80

250

200

100

50

150

10 20

DBC

Recovery

2 4 6 8 12 14 16 18

DBC（mg/mL-resin）

Recovery（％）

initial

10th

20th

1 2 3

min10 20 30 40 50

Number of CIP

Separation of standard proteins

1. Ribonuclease A2. Cytochrome c3. Lysozyme

Purification of IgY from egg yolk extract

Egg yolk antibody (IgY) can be isolated with high purity more than 99% by two chromatographic purification steps, which consist of a capture step by ion exchange chromatography on YMC-BioPro Q75 and a polishing step by size exclusion chromatography on YMC-Pack Diol-200.

min

mV

0 5 10 15 20 25 30

0

25

50

75

100

YMC-Pack Diol-200 5 µm, 300×8.0 mmI.D.Sample : Fraction from capture purification by IEC

Polishing by size exclusion chromatography (SEC)

Analysis of purified fraction


IgY

min

mAU

0 5 10 15 20

0.0

2.5

5.0

7.5

97.2 kDa

66.4 kDa

45.0 kDa

29.0 kDa

20.1 kDa14.3 kDa

Marker

StandardIgY

Non-reduced SDS-PAGEIEC

fractionSEC

fractionCrudeextract

purity >99%

IgY

SEC

Fraction from polishing step by SEC

Eluent ： 0.1 M KH2PO4-K2HPO4 (pH 6.9) containing 0.2 M NaCl

Flow rate ：0.7 mL/minTemperature ：ambientDetection ：UV at 280 nmInjection ：1 mL (ca. 0.45 mg IgY)

min

mV

0 10 20 30 40 50 60

0

100

200

300

400

500

YMC-BioPro Q75 75 µm, 50×4.6 mmI.D.Sample : Crude extract from egg yolk*　

Capture purification by ion exchange chromatography (IEC)

Polishing step

Eluent ： A) 20 mM Tris-HCl (pH 8.1) B) 20 mM Tris-HCl (pH 8.1) containing 0.5 M NaCl 10% B (0-15 min), 30% B (15-30 min), 90% B (30-40 min)

Flow rate ：0.5 mL/min (180 cm/hr)Temperature ：ambientDetection ：UV at 280 nmInjection ：1 mL (ca. 20 mg Protein)

＊Courtesy of Pharma Foods International Co., Ltd.

Packing material Particle size（µm）

Productnumber

Volume50 mL 250 mL 1 L 5 L 25 L

YMC-BioPro Q30 30 QAA0S30 ＊＊＊＊＊YMC-BioPro S30 30 SPA0S30 ＊＊＊＊＊YMC-BioPro Q75 75 QAA0S75 ＊＊＊＊＊YMC-BioPro S75 75 SPA0S75 ＊＊＊＊＊

Ordering Information

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Silica gel SEC

YMC-Pack Diol● 5 µm silica-based column with high mechanical stability● Low-cost size exclusion chromatography (SEC) column● Useful for molecular weight determination of proteins and sugars

S i l i c a - b a s e d s i z e e x c l u s i o n c h r o m a t o g r a p h y ( S E C ) c o l u m nYMC-Pack Diol is a size exclusion chromatography column based dihydroxypropyl-bonded silica, and available in four different pore sizes. Diol-120, 200, and 300 are suitable for separation or molecular weight

determination of proteins with molecular weights of 5,000 to several hundred thousand. Diol-60 is the most suitable for separation of peptides or oligosaccharides whose molecular weights are 10,000 or less.

■Particle size：5 µm■Pore size：60, 120, 200, 300 Å■Usable pH range：5.0 〜 7.5

Column ： YMC-Pack Diol 300×8.0 mmI.D.

Eluent ：0.1 M KH2PO4-K2HPO4 (pH 7.0) containing 0.2 M NaClFlow rate ：0.5 mL/minTemperature ：25 ℃Detection ：UV at 280 nm

Calibration curves of various proteins for three different pore sizes

5 131197

Elution volume (mL)

MW

104

105

106

Diol-300

Diol-200Diol-120

1

11

14 15

9

87

65

43

2

10

12

16

13

Diol-120, Diol-200 and Diol-300 are suitable for the separation or molecular weight determination of proteins with molecular weights of 5,000 to several hundred thousand.

MW 1. IgM 900,000 2. Thyroglobulin 670,000 3. IgA 390,000 4. Fibrinogen 340,000 5. γ-Globulin 158,000 6. IgG 150,000 7. Transferrin 75,000 8. HSA (human serum albumin) 66,000 9. α1-Antitrypsin 50,00010. Ovalbumin 45,00011. Carbonic anhydrase 30,00012. Trypsin inhibitor 20,10013. Myoglobin 17,00014. α-Lactalbumin 14,10015. Ribonuclease A 13,70016. Cytochrome c 12,400

Separation for standard protein markers

10 20 30

1

54

32

0 min

Diol-120

For molecular weight 10,000 to 500,000 compounds, Diol-200 is suitable for the separation.

Column ： YMC-Pack Diol 500×8.0 mmI.D.

Eluent ： 0.1 M KH2PO4-K2HPO4 (pH 7.0) containing 0.2 M NaCl

Flow rate ：0.7 mL/minTemperature ：ambientDetection ：UV at 280 nm

MW1. Glutamate dehydrogenase 290,0002. Lactate dehydrogenase 142,0003. Enolase 67,0004. Adenylate kinase 32,0005. Cytochrome c 12,400

1

54

32

10 20 300 40min

Diol-200

1 543

2

10 20 300 40 min

Diol-300

Specifications

Column Base Functional groupPore size (Å)

Particle size

(µm)

Usable pH

rangeCharacteristics

Diol-60

Silica gel Dihydroxypropyl

60

5 5 〜 7.5

For molecular weight below 10,000

Diol-120 120 For molecular weight 5,000 to 100,000

Diol-200 200 For molecular weight 10,000 to ca. 500,000

Diol-300 300 For molecular weight ca. 50,000 to 1,000,000

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Separation for molecular weight below 10,000 peptides

Diol-60

10 20 30 40 min

12

34

0

For molecular weight below 10,000 peptides, Diol-60 is suitable for the separation.

MW1. Insulin (Bovine) 5,7002. Neurotensin 1,6723. Angiotensin II 1,0404. Glycine 75

Column ： YMC-Pack Diol-60 500×8.0 mmI.D.

Eluent ： 0.1 M KH2PO4-K2HPO4

(pH 7.0) containing 0.2 M NaCl/ acetonitrile (70/30)


MW 1. Myoglobin 17,000 2. Ribonuclease A 13,700 3. Cytochrome c 12,400 4. Insulin (Bovine) 5,700 5. Insulin B chain 3,496 6. α-Mating factor 1,684 7. Neurotensin 1,672 8. Angiotensin I 1,296 9. CCK-Octapeptide 1,14310. Bradykinin 1,06011. Angiotensin II 1,04012. Oxytocin 1,00713. Angiotensin III 93114. Met-Enkephalin 57315. Leu-Enkephalin 55516. Tetraglycine 24617. Glycine 75

10 15 20

102

103

104

MW

Elution volume (mL)

1 2

3

4

5

7 68

911 13 10

12

1514

16

17

Separation of oligo- and polysaccharide

Diol-200

10 20 min

1

2

34

0

For separation or molecular weight determination of water-soluble oligo- and polysaccharides, Diol-60, Diol-120, Diol-200, and Diol-300 are useful individually or in combination.

Column ：YMC-Pack Diol, 500×8.0 mmI.D.Eluent ：waterFlow rate ：1.0 mL/minTemperature ：ambientDetection ：RI

MW 1. Pullulan (P-800) 853,000 2. Pullulan (P-400) 380,000 3. Pullulan (P-200) 186,000 4. Pullulan (P-100) 100,000 5. Pullulan (P-50) 48,000 6. Pullulan (P-20) 23,700 7. Pullulan (P-10) 12,200 8. Pullulan (P-5) 5,800 9. Maltopentadecaose (G15) 2,44810. Maltoundecaose (G11) 1,80011. Maltoheptaose (G7) 1,15212. Maltopentaose (G5) 82413. Maltotriose (G3) 50414. Maltose (G2) 34215. Glucose (G1) 180

MW1. P-800 853,0002. P-50 48,0003. P-20 23,7004. P-5 5,800

10 15 20 25

102

103

104

105

106

MW

Elution volume (mL)

1011

1213

1415

9

8

7

6

5

43

2

1

Diol-60

Diol-300

Diol-200

Diol-120

Analytical columns（Stainless columns）Packing material

Pore size（Å）


Diol-60 60 4.6×300 DL06S05-3046WT8.0×300 DL06S05-3008WT8.0×500 DL06S05-5008WT




For preparative columns with an inner diameter of 20 mm or more, see page 153.

Guard columns（Stainless columns）Packing material

Pore size（Å）


Diol-60 60 8.0×30 DL06S05-0308WTG

Diol-120 120 8.0×30 DL12S05-0308WTG

Diol-200 200 8.0×30 DL20S05-0308WTG

Diol-300 300 8.0×30 DL30S05-0308WTG

Analytical columns（Glass columns）Diol-60 60 8.0×300 DL06S05-3008FG

8.0×500 DL06S05-5008FG

Diol-120 120 8.0×300 DL12S05-3008FG8.0×500 DL12S05-5008FG

Diol-200 200 8.0×300 DL20S05-3008FG8.0×500 DL20S05-5008FG

Diol-300 300 8.0×300 DL30S05-3008FG8.0×500 DL30S05-5008FG


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Reversed-phase columns

Reversed-phase columns for BIO-MACRO molecular separations● Abundant gel types● Excellent peak shapes● High resolution

Reve r s ed - p ha se co l umns f o r p ep t i d e s , p r o t e i n s a nd n uc l e i c a c i d sFor BIO-MACRO molecular separations, YMC has many choices for reversed-phase HPLC.A listing of the most popular column is found below.

Specifications　For products other than below listed, see the "Reversed-phase C18 columns (ODS)" and "Reversed-phase columns (other than ODS)" sections.

Separation for molecular weight below 5,000 (pore size 80 Å, 120 Å)

Column Base Functional group

Pore size(Å)

Particle size(µm)

Carbon content (% )

Usable pH range Characteristics

Triart C18 Hybrid silica C18 120 2, 3, 5 20 1.0 〜12.0 ・Excellent chemical durability

・Superior peak shapes in every condition

Pro

ser

ies

Pro C18

Silica gel

C18

120 2*, 3, 5,10 162.0 〜 8.0

・ Processed with YMC's advanced endcapping technology・ Superior separation for basic compounds

Hydrosphere C18 120 2*, 3, 5 12 ・Superior separation for hydrophilic compounds

・Can be used with 100% water mobile phase

Pro C18 RS 80 3, 5 22 1.0 〜10.0

・Highly durable ODS・ Superior separation for basic compounds

and hydrophobic compounds

YMC-

Pack

serie

s ODS-A 120 3, 5,10,15, 20, 50

172.0 〜 7.5

・Currently in use worldwide・Available from analytical to preparative

ODS-AQ 120 14 ・Superior separation for hydrophilic compoundsPh Phenyl 120 3, 5,10,15, 20 9 ・ Reversed-phase packing material with π electronsPolymerC18 Polymer C18 − 6,10 − 2.0 〜13.0 ・Polymer based ODS

* Materials for 2 µm is YMC-UltraHT products.

Separation for molecular weight 5,000 - 100,000 (pore size 200 Å, 300 Å)

Column Base Functional group

Pore size(Å)

Particle size(µm)

Carbon content (% )

Usable pH range Characteristics

Wid

e po

re c

olum

n

ODS-A

Silica gel

C18200 5,10,

15, 2012

2.0 〜 7.5

・ODS with wide pore size・For separation of peptides and proteins300 7

ODS-AQ C18 200 5,10,15 10 ・Low carbon ODS

C8 C8200 5,10,

15, 207 ・C8 with wide pore size

・For separation of relatively highly hydrophobic compounds300 4

C4 C4200 5,10,

15, 205 ・C4 with wide pore size

・Superior separation for proteins300 3

CN cyano propyl 300 5 3 ・CN with wide pore size

・Unique selectivity due to cyano group

YMCbasic C8 200 3, 5 7 ・ Superior separation for proteins and peptides, especially for insulin

PROTEIN-RP − 200 5 4 1.5 〜 7.5 ・ Specialized column with excellent acid resistance for separation of proteins and peptides

How to select reversed-phase columnsTo separate proteins or peptides, select columns based on the molecular weight of the compounds to be separated is important. As shown in the table on the right, the C18 column with 120 Å pore size is generally suitable for small peptides up to MW 5,000. In the case of large peptides or small proteins up to MW 20,000, the C8 column with 200 Å pore size often gives the best column efficiency. Furthermore, most of proteins are eluted effectively by the C4 column with 300 Å. Separation may also be influenced by the hydrophobicity of the analyte and the type of the functional group as well as molecular weight. If the sufficient separation is not achieved with columns marked with a double circle, perform optimization as indicated by the arrows shown in the table. In addition to columns C18, C8, and C4 shown in the table, PROTEIN-RP and CN type columns with different selectivity are also useful.

Molecular weightof sample

Functionalgroup

Pore sizeC18 C8 C4

120 Å ◎ ○ ○

200 Å ○ ◎ ○

300 Å ○ ○ ◎

5,000

20,000

100,000

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Separation of peptides (MW 573 - 3,465)

Generally, the conventional C18 column with 120 Å pore size is suitable for analysis of small peptides up to 5,000 in molecular weight. Especially Pro series ODS columns, which are processed with advanced endcapping technology, are ideal for separation of basic peptides. As shown in the above, Hydrosphere C18, a Pro series column, exhibits excellent separations and superior peak shapes of basic peptides (peak 1 and 7), in contrast to the commercial ODS column for hydrophilic compounds, Brand E2.

Eluent ： A) water/TFA (100/0.1) B) acetonitrile/TFA (100/0.1) 20-40% B (0-15 min), 40% B (15-20 min)

Flow rate ：1.0 mL/minTemperature ：37 ℃Detection ：UV at 220 nm

1. BAM-12P (MW 1,425)2. [D-Ala2,Met5]-Enkephalinamide (MW 587)3. α-Endorphin (MW 1,746)4. Met-Enkephalin (MW 573)5. [D-Ala2,Met5]-Enkephalin (MW 588)6. γ-Endorphin (MW 1,899)7. β-Endorphin (MW 3,465)

Hydrosphere C18（120 Å）5 µm,150×4.6 mmI.D.Hydrosphere C18（120 Å）5 µm,150×4.6 mmI.D.

1

23

45

6

7

0 5 10 15 20 min

A000308B

Excellent peak shapes for basic peptides

Brand E2（100 Å）5 µm,150×4.6 mmI.D.(ODS column for hydrophilic compounds)

Brand E2（100 Å）5 µm,150×4.6 mmI.D.(ODS column for hydrophilic compounds)

1

2 34

5 6

7

0 5 10 15 20 min

A000313D

Separation of peptides and proteins (MW 4,300 - 17,000)

For proteins and peptides with molecular weight of 4,300 to 17,000, separation characteristics are compared using columns with different pore size and functional group. In accordance with the table on the previous page, the suitable column is C8, 200 Å for groups of compounds with a molecular weight within this range. If either pore size or functional group of the packing material is not optimized, peak broadening and poor resolution are observed. By using the most suitable column (C8, 200 Å) for the target compounds, sharp peak shapes and excellent separation are achieved.

Column ：5 µm, 150×4.6 mmI.D.Eluent ： A) water/TFA (100/0.1)

B) acetonitrile/TFA (100/0.1) 25-60% B (0-20 min)


1. Cytochrome c (MW 12,400)2. Insulin (Bovine) (MW 5,700)3. Amyloid β-protein (MW 4,300)4. Lysozyme (MW 14,300)5. α-Lactalbumin (MW 14,100)6. Myoglobin (MW 17,000)

C8, 300 Å

C8, 120 Å

04073006.D

min0 5 10 15

04080204.D

1 2

56

4

3

12

35

64

Pore size

Comparison of separation on columns with different pore size and functional group

C4, 200 Å

C18, 200 Å

min0 5 10 1504073002.D

04072910.D

12

35

64

1,2

3 5

64

Functional group

1

2

35

64

min0 5 10 15

04082002.D

C8, 200 Å

Optimized combination of pore size and functional group

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Reversed-phase columns for BIO-MACRO molecular separations

Separation of proteins (MW 66,000 - 96,000)

Gradient elution of water and acetonitrile containing TFA are often employed in an analysis of proteins and peptides. In some cases, addition of a “third solvent” is effective for change in selectivity and separation. The above example shows the resolution between high-molecular weight proteins (peak 1 and 2) is improved by adding 2-propanol into the standard mobile phase of acetonitrile/water/TFA.

Column ： YMC-Pack C4 (5 µm, 300 Å) 150×4.6 mmI.D.

Eluent ： 30-75% B (0-15 min), 75% B (15-20 min)


1. BSA (MW 66,000)2. Conalbumin (MW 77,000)3. Lipoxidase (MW 96,000)

Optimization of eluent conditions (C4, 300 Å)

Separation characteristics of proteins with molecular weight of 66,000 to 96,000 are compared using columns with different pore size and functional group. The columns with smaller pore size, which have the same C4 functional groups, provide broader peak shapes and poor separations. In comparison among the 300 Å pore columns with different functional groups, the longer alkyl chain such as C18 and C8 results in poor resolution. It is important to choose optimal pore size and functional group depending on molecular weight of proteins for better peak shapes and resolutions. Proteins with molecular weight of 20,000 to 100,000 are separated effectively by the C4 column with 300 Å pore size.

1. BSA (MW 66,000)2. Conalbumin (MW 77,000)3. Lipoxidase (MW 96,000)

Comparison of separation on columns with different pore size and functional group

Column ：5 µm, 150×4.6 mmI.D.Eluent ： A) water/TFA (100/0.1)

B) acetonitrile/2-propanol/TFA（50/50/0.1） 30-75% B (0-15 min), 75% B (15-20 min)


1

3

2

A） water/TFA（100/0.1）B） acetonitrile/TFA（100/0.1）

05011405.D

0 5 10 15 min

3

1

2

A） water/TFA（100/0.1）B） acetonitrile/2-propanol/TFA（50/50/0.1）

04093007.D

0 5 10 15 min

2-propanoladdition

12

3

12 3

C4, 200 Å

C4, 120 Å

0 4 8 12 16 min

Pore size

05011714.D

05011711.D

1

2

3

C4, 300 Å

04093007.D

12 16 min840

1 23

1 23

0 4 8 12 16 min

C8, 300 Å

C18, 300 Å

Functional group

05011708.D

05011802.D

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Separation of nucleic acid bases and nucleotides

0S991029A

10 20 minA000922A0 5 10 15 20 min

Nucleic acid bases Nucleotides

Hydrosphere C18 is designed for separation of hydrophilic compounds and can be used with 100% aqueous mobile phase. Hydrosphere C18 is favorable to the analyses of highly polar compounds such as nucleic acid bases and nucleotides.

Column ： Hydrosphere C18 (5 µm, 120 Å) 150×4.6 mmI.D.

Eluent ：20 mM KH2PO4-K2HPO4 (pH 6.9)Flow rate ：1.0 mL/minTemperature ：37 ℃Detection ：UV at 254 nm

Column ： Hydrosphere C18 (5 µm, 120 Å) 150×4.6 mmI.D.　

Eluent ：100 mM KH2PO4-K2HPO4 (pH 5.5)Flow rate　：1.0 mL/minTemperature ：30 ℃Detection ：UV at 260 nm

In analytical scale, many impurities could be separated from the target compound by one-nucleotide difference on Hydrosphere C18. Even in purification scale, YMC-Actus gave superior separation and recovery.YMC-Actus Hydrosphere C18 is useful for purification of hydrophilic compounds such as oligonucleotides, organic acids, oligosaccharides and glycosides.

Eluent ： A）10 mM DBAA*（pH 6.0）/methanol（60/40） B）10 mM DBAA*（pH 6.0）/methanol（20/80） 10-35% B（0-30 min）

Temperature ：ambientDetection ：UV at 269 nmSample ：synthetic oligonucleotide（100 µM）* di-n-butylammonium acetate

5'-CCGCTCGAGCTAAAAAAAGCCTGTGTTACC-3'

Purification of highly polar compounds　－oligonucleotide－Crude synthetic 30 mer oligonucleotide

Hydrosphere C18 5 µm,50×4.6 mmI.D.

YMC-Actus Hydrosphere C18 5 µm,50×20 mmI.D.

Brand I1 5 µm,50×4.6 mmI.D.

Brand I1 5 µm,50×19 mmI.D.

Analysis Purification1.0 mL/min, 5 µL injection 19 mL/min, 100 µL injection

30 mer

30 mer

Impurities

Recovery 56%■ purity>99%

0 10 20 30 min 20 22.5 25 27.5 min

Recovery 35%■ purity>99%

Impurities

7.5 10 12.50 10 20 30 min min

Separation of oligonucleotides

0 5 10 15 20 25 30 min

T2 T10

T20

T2-20

T10

T20

T20T10

T2

T2

J050214A

J050209B

J050210E

J050209A

Hydrosphere C18, 10 mM TEAA

Pro C18, 10 mM TEAA

Brand I1, 10 mM TEAA

Brand I1, 100 mM TEAA Hydrosphere C18 shows strong retention and excellent resolution of oligonucleotides even at a low concentration of triethylamine compared to ordinary C18 columns.

Column ：5 µm, 150×4.6 mmI.D.Eluent ： A) 10 mM or 100 mM TEAA* (pH 6.0)

B) 10 mM or 100 mM TEAA* (pH 6.0)/acetonitrile (80/20) 55-61% B (0-30 min)

Flow rate ：1.0 mL/minTemperature ：35 ℃Detection ：UV at 269 nm* triethylammonium acetate

Oligonucleotides d（pT）2-20

1. 5'-UTP 2. 5'-UDP 3. 5'-CMP 4. 5'-UMP 5. 5'-dCMP 6. 5'-GMP 7. 5'-ATP 8. 5'-IMP 9. 5'-ADP10. 5'-TMP11. 5'-AMP

See P138-141 for details of YMC-Actus series.

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YMC-Pack ODS-A

Analytical columnsParticle size（µm）

Pore size（Å）


S-5 200 4.6×150 AA20S05-1546WT4.6×250 AA20S05-2546WT

300 1.0×150 AA30S05-1501WT1.0×250 AA30S05-2501WT1.5×150 AA30S05-15P5WT1.5×250 AA30S05-25P5WT2.0× 75 AA30S05-L502WT2.0×150 AA30S05-1502WT2.0×250 AA30S05-2502WT4.6× 75 AA30S05-L546WT4.6×100 AA30S05-1046WT4.6×150 AA30S05-1546WT4.6×250 AA30S05-2546WT4.6×300 AA30S05-3046WT6.0×100 AA30S05-1006WT6.0×150 AA30S05-1506WT6.0×250 AA30S05-2506WT6.0×300 AA30S05-3006WT

S-10 300 4.6×100 AA30S11-1046WT4.6×150 AA30S11-1546WT4.6×250 AA30S11-2546WT4.6×300 AA30S11-3046WT6.0×100 AA30S11-1006WT6.0×150 AA30S11-1506WT6.0×250 AA30S11-2506WT6.0×300 AA30S11-3006WT

Semi-preparative columnsS-5 200 20×150 AA20S05-1520WT

20×250 AA20S05-2520WT

300 10×150 AA30S05-1510WT10×250 AA30S05-2510WT10×300 AA30S05-3010WT20×150 AA30S05-1520WT20×250 AA30S05-2520WT

S-10 200 20×150 AA20S11-1520WT20×250 AA20S11-2520WT

300 10×150 AA30S11-1510WT10×250 AA30S11-2510WT10×300 AA30S11-3010WT20×150 AA30S11-1520WT20×250 AA30S11-2520WT

See P145 for preparative columns other than those listed above.

Columns shown in above are only wide pore size columns. Details of pore sizes for 80 Å and 120 Å columns, see the pages of reversed-phase columns.

Guard cartridge columns (with inner diameter 1.0, 1.5, or 2.0 mm: 2/pack, 4.0 mm: 3/pack)Particle size（µm）

Pore size（Å）


S-5 200 4.0×23 AA20S05-G304CC

300 1.0×10 AA30S05-0101CC1.5×10 AA30S05-01P5CC2.0×10 AA30S05-0102CC4.0×23 AA30S05-G304CC

A guard cartridge holder will need to be purchased separately before using this product for the first time.

Cartridge holderSemi-micro guard cartridge holder(inner diameter: 1.0, 1.5, or 2.0 mm)

XPCHSMW

Cartridge holder(inner diameter: 4.0 mm)

XPCHW

YMC guard cartridge packS-5 200 4.0×23 AA20S05-G304CCPK

300 4.0×23 AA30S05-G304CCPK

A guard cartridge pack includes a cartridge holder (1 set), a guard cartridge column, and a column coupler (produced by PEEK).

Guard columnsS-5 200 4.0×10 AA20S05-0104WFG

20×50 AA20S05-0520WTG

300 4.0×10 AA30S05-0104WFG5.0×10 AA30S05-0105WFG10×30 AA30S05-0310WTG20×50 AA30S05-0520WTG

S-10 200 20×50 AA20S11-0520WTG

300 4.0×10 AA30S11-0104WFG5.0×10 AA30S11-0105WFG10×30 AA30S11-0310WTG20×50 AA30S11-0520WTG


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YMC-Pack ODS-AQ


Pore size（Å）


S-5 200 4.6×150 AQ20S05-1546WT4.6×250 AQ20S05-2546WT

Semi-preparative columnsS-5 200 20×150 AQ20S05-1520WT

20×250 AQ20S05-2520WT

S-10 200 20×150 AQ20S11-1520WT20×250 AQ20S11-2520WT


Guard cartridge columns (three-pack)S-5 200 4.0×23 AQ20S05-G304CC

A cartridge holder will need to be purchased separately before using this product for the first time.

Cartridge holderCartridge holder(inner diameter: 4.0 mm)

XPCHW

YMC guard cartridge packS-5 200 4.0×23 AQ20S05-G304CCPK

A guard cartridge pack includes a cartridge holder (1 set), a guard cartridge column, and a column coupler (produced by PEEK).

Guard columnsS-5 200 4.0×10 AQ20S05-0104WFG

20×50 AQ20S05-0520WTG

S-10 200 20×50 AQ20S11-0520WTG


YMC-Pack CN


Pore size（Å）


S-5 300 1.0×150 CN30S05-1501WT1.0×250 CN30S05-2501WT1.5×150 CN30S05-15P5WT1.5×250 CN30S05-25P5WT2.0×150 CN30S05-1502WT2.0×250 CN30S05-2502WT4.6× 75 CN30S05-L546WT4.6×100 CN30S05-1046WT4.6×150 CN30S05-1546WT4.6×250 CN30S05-2546WT4.6×300 CN30S05-3046WT6.0×100 CN30S05-1006WT6.0×150 CN30S05-1506WT6.0×250 CN30S05-2506WT6.0×300 CN30S05-3006WT

Semi-preparative columnsS-5 300 20×150 CN30S05-1520WT

20×250 CN30S05-2520WTSee P151 for preparative columns other than those listed above.

Guard cartridge columns (three-pack)S-5 300 4.0×23 CN30S05-G304CC



XPCHW

Guard columnsS-5 300 4.0×10 CN30S05-0104WFG

5.0×10 CN30S05-0105WFG10×30 CN30S05-0310WTG20×50 CN30S05-0520WTG


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YMC-Pack C8


Pore size（Å）


S-5 200 4.6×150 OC20S05-1546WT4.6×250 OC20S05-2546WT

300 1.0×150 OC30S05-1501WT1.0×250 OC30S05-2501WT1.5×150 OC30S05-15P5WT1.5×250 OC30S05-25P5WT2.0×150 OC30S05-1502WT2.0×250 OC30S05-2502WT4.6× 75 OC30S05-L546WT4.6×100 OC30S05-1046WT4.6×150 OC30S05-1546WT4.6×250 OC30S05-2546WT4.6×300 OC30S05-3046WT6.0×100 OC30S05-1006WT6.0×150 OC30S05-1506WT6.0×250 OC30S05-2506WT6.0×300 OC30S05-3006WT

S-10 300 4.6×100 OC30S11-1046WT4.6×150 OC30S11-1546WT4.6×250 OC30S11-2546WT4.6×300 OC30S11-3046WT6.0×100 OC30S11-1006WT6.0×150 OC30S11-1506WT6.0×250 OC30S11-2506WT6.0×300 OC30S11-3006WT

Semi-preparative columnsS-5 200 20×150 OC20S05-1520WT

20×250 OC20S05-2520WT

300 10×150 OC30S05-1510WT10×250 OC30S05-2510WT10×300 OC30S05-3010WT20×150 OC30S05-1520WT20×250 OC30S05-2520WT

S-10 200 20×150 OC20S11-1520WT20×250 OC20S11-2520WT

300 10×150 OC30S11-1510WT10×250 OC30S11-2510WT10×300 OC30S11-3010WT20×150 OC30S11-1520WT20×250 OC30S11-2520WT




Pore size（Å）


S-5 200 4.0×23 OC20S05-G304CC

300 1.0×10 OC30S05-0101CC1.5×10 OC30S05-01P5CC2.0×10 OC30S05-0102CC4.0×23 OC30S05-G304CC



XPCHSMW


XPCHW

Guard columnsS-5 200 4.0×10 OC20S05-0104WFG

20×50 OC20S05-0520WTG

300 4.0×10 OC30S05-0104WFG5.0×10 OC30S05-0105WFG10×30 OC30S05-0310WTG20×50 OC30S05-0520WTG

S-10 200 20×50 OC20S11-0520WTG

300 4.0×10 OC30S11-0104WFG5.0×10 OC30S11-0105WFG10×30 OC30S11-0310WTG20×50 OC30S11-0520WTG


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YMC-Pack C4


Pore size（Å）


S-5 200 4.6×150 BU20S05-1546WT4.6×250 BU20S05-2546WT

300 1.0×150 BU30S05-1501WT1.0×250 BU30S05-2501WT1.5×150 BU30S05-15P5WT1.5×250 BU30S05-25P5WT2.0×150 BU30S05-1502WT2.0×250 BU30S05-2502WT4.6× 75 BU30S05-L546WT4.6×100 BU30S05-1046WT4.6×150 BU30S05-1546WT4.6×250 BU30S05-2546WT4.6×300 BU30S05-3046WT6.0×100 BU30S05-1006WT6.0×150 BU30S05-1506WT6.0×250 BU30S05-2506WT6.0×300 BU30S05-3006WT

S-10 300 4.6×100 BU30S11-1046WT4.6×150 BU30S11-1546WT4.6×250 BU30S11-2546WT4.6×300 BU30S11-3046WT6.0×100 BU30S11-1006WT6.0×150 BU30S11-1506WT6.0×250 BU30S11-2506WT6.0×300 BU30S11-3006WT

Semi-preparative columnsS-5 200 20×150 BU20S05-1520WT

20×250 BU20S05-2520WT

300 10×150 BU30S05-1510WT10×250 BU30S05-2510WT10×300 BU30S05-3010WT20×150 BU30S05-1520WT20×250 BU30S05-2520WT

S-10 200 20×150 BU20S11-1520WT20×250 BU20S11-2520WT

300 10×150 BU30S11-1510WT10×250 BU30S11-2510WT10×300 BU30S11-3010WT20×150 BU30S11-1520WT20×250 BU30S11-2520WT




Pore size（Å）


S-5 200 4.0×23 BU20S05-G304CC

300 1.0×10 BU30S05-0101CC1.5×10 BU30S05-01P5CC2.0×10 BU30S05-0102CC4.0×23 BU30S05-G304CC



XPCHSMW


XPCHW

Guard columnsS-5 200 4.0×10 BU20S05-0104WFG

20×50 BU20S05-0520WTG

300 4.0×10 BU30S05-0104WFG5.0×10 BU30S05-0105WFG10×30 BU30S05-0310WTG20×50 BU30S05-0520WTG

S-10 200 20×50 BU20S11-0520WTG

300 4.0×10 BU30S11-0104WFG5.0×10 BU30S11-0105WFG10×30 BU30S11-0310WTG20×50 BU30S11-0520WTG

See P76 for YMCbasic.See P122, 123 for YMC-Pack PROTEIN-RP.


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YMC-Pack PROTEIN-RP● Improved recovery of proteins or peptides● Improved durability when used with TFA solution● Enables elution of high molecular weight proteins

R e v e r s e d - p h a s e c o l u m n f o r s e p a r a t i o n o f p r o t e i n s o r p e p t i d e sYMC-Pack PROTEIN-RP is a reversed-phase column utilizing a silica gel base. It contains a stationary phase, specifically designed for separation of proteins or peptides. Problems that are associated with

conventional reversed-phase columns with short alkyl chain lengths are minimized.Robust column lifetime and excellent recovery of hydrophobic proteins are typically possible on this phase.

■Carbon content：4％■Usable pH range：1.5 〜 7.5

Recovery of various proteins and peptides is shown.YMC-Pack PROTEIN-RP gives greater recovery than competitive C4 columns.

Improved recovery of proteins or peptidesRecovery of proteins or peptides

SampleRecovery（％）

YMC-PackPROTEIN-RP

C4 column,company A

C4 column,company B

Ribonuclease A 93 95 86Cytochrome c 94 89 95Lysozyme 98 93 76Myoglobin 97 85 93BSA 94 92 86Ovalbumin 90 73 88Transferrin 94 98 88Insulin(bovine) 97 73 92Insulin B chain 82 76 70α-Mating factor 93 82 81Leu-Enkephalin 92 84 91Gly-Gly-Gly-Gly 95 86 70

Improved durability when used with TFA solution

110

100

90

80

70

60

50

40

30

20

10

0100 200 300 400

C4 column, Company B

C4 column, Company A

（Peak split）

YMC-Pack PROTEIN-RP

×

Solvent flow in hours

0.1％TFA condition

%

retention

of

diphenyl

The left graph shows the test result of the stability of the stationary phase with 0.1% aqueous TFA. Retention of diphenyl on C4 columns manufactured by other companies greatly decreases as time passes. This is caused by detachment of butyl groups from the packing material due to acid hydrolysis.Retention of diphenyl on YMC-Pack PROTEIN-RP is shown to be stable after 500 hours of mobile phase flow.

〈Prior to flow〉1

2

3

4

0 5 10 min

Theoretical plate number (4) = 14,300

〈After 500 hours〉

1

2

3

4

0 5 10 min

Theoretical plate number (4) = 13,800

1. Uracil2. Benzene3. Naphthalene4. Diphenyl

〈Flow conditions〉Eluent ：water/TFA (100/0.1)Flow rate ：1.0 mL/minTemperature ：ambient

〈Measurement conditions〉Column ：YMC-Pack PROTEIN-RP 　250×4.6 mmI.D.Eluent ：acetonitrile/water (40/60)Flow rate ：1.0 mL/minTemperature ：30 ℃Detection ：UV at 254 nm, 0.32 AUFS

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0 10 20 30 min

12

3

Proteins1. Cytochrome c （MW 12,400）2. BSA （MW 66,000）3. Ferritin （MW 440,000）

Column ：YMC-Pack PROTEIN-RP 　250×4.6 mmI.D.Eluent ： A) water/TFA (100/0.1)

B) acetonitrile/TFA (100/0.1) 　30-90％ B (0-45 min, linear)Flow rate ：1.0 mL/minTemperature ：ambientDetection ：UV at 280 nm, 0.04 AUFS

A p p l i c a t i o n

1

23 4 5

67

8

9

1011

12

13

403020100 min

Proteins and peptides 1. Met-Enkephalin （MW 573） 2. Leu-Enkephalin （MW 555） 3. Oxytocin （MW 1,007） 4. Bradykinin （MW 1,060） 5. Angiotensin I （MW 1,296） 6. Ribonuclease A （MW 13,700） 7. α-Mating factor （MW 1,684） 8. Insulin（bovine）（MW 5,700） 9. Cytochrome c （MW 12,400） 10. Lysozyme （MW 14,400） 11. BSA （MW 66,000） 12. β-Lactoglobulin （MW 18,400） 13. Ovalbumin （MW 45,000）

Column ：YMC-Pack PROTEIN-RP 　250×4.6 mmI.D.Eluent ： A) water/TFA (100/0.1)

B) acetonitrile/TFA (100/0.1) 　10-90％ B (0-60 min, linear)Flow rate ：1.0 mL/minTemperature ：ambientDetection ：UV at 220 nm, 0.32 AUFS

A p p l i c a t i o n

Analytical columnsParticle size

（µm）Column size

inner diameter×length（mm） Product number

S-5 2.0×150 PR99S05-1502WT2.0×250 PR99S05-2502WT4.6×150 PR99S05-1546WT4.6×250 PR99S05-2546WT

Semi-preparative columnsS-5 10×250 PR99S05-2510WT

20×150 PR99S05-1520WT20×250 PR99S05-2520WT

Guard cartridge columns (three-pack)S-5 4.0×23 PR99S05-G304CC



XPCHW

Guard columnsS-5 4.0×10 PR99S05-0104WFG

10×30 PR99S05-0310WTG20×50 PR99S05-0520WTG


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Normal-phase columns

YMC-Pack Diol-NP● Silica gel bonded with dihydroxypropyl group● Two pore sizes of 120 Å and 60 Å are available● Normal-phase separation using non-polar

solvents

● Useful for hydrophilic interaction chromatography (HILIC)

● Different separation characteristics from bare silica gel

Diol column for non-aqueous Normal-Phase (NP) and Hydrophilic Interaction Chromatography (HILIC) YMC-Pack Diol-NP shows retention behavior of normal-phase chromatography when it is used with low-polarity mobile phases. Hydroxy groups on the surface of the stationary phase act as polar groups. YMC-Pack Diol-NP is as widely applicable to normal-phase separation as silica

gel. It is also useful in cases where separation optimization is difficult to achieve using bare silica gel. In addition, YMC-Pack Diol-NP is usuable as a HILIC mode column when it is used with a mobile phase consisted of a high concentration of organic solvent in water or an aqueous buffer.

■Pore size：120 Å, 60 Å■Usable pH range：2.0 〜 7.5

Separation of highly hydrophilic compounds in HILIC mode

0 105 min15

1

5

4

32

6

R090116P

YMC-Pack Diol-NP shows superior performance as a HILIC mode column when it is used with a mobile phase consisted of a mixture of organic solvent/aqueous solution.

Column ： YMC-Pack Diol-NP (5 µm, 120 Å) 150×2.0 mmI.D.

Eluent ：water/acetonitrile (10/90) containing 10 mM CH3COONH4


1. Uracil 2. Uridine 3. Adenosine 4. Cytosine 5. Cytidine 6. Guanosine

NH

HN

O

O

N

HN

OH

O

OH

HOH2CO

O

N

N

N

N

OH

O

OH

HOH2C

NH2

NH

N

O

NH2

N

N

OH

O

OH

HOH2CO

NH2

HN

N

N

N

O

OH

O

OH

HOH2C

H2N

Nucleic acid bases and Nucleosides

Separation of hydrophilic compounds which are hardly retained on reversed-phase column

0 2 4 6 min

A991115E

YMC-Pack Diol-NP（5 µm, 120 Å） 50×4.6 mmI.D.

1

2

3

45

Comparison of peptide separation on YMC-Pack Diol-NP and YMC-Pack ODS-A is shown. YMC-Pack Diol-NP offers favorable resolution of between peak 1 and peak 2, highly hydrophilic peptides which cannot be separated in reversed-phase mode with ODS column. An interaction between hydroxyl groups on the Diol-NP stationary phase and highly hydrophilic compounds provides enhanced retention and superior resolution.

Eluent ： A) acetonitrile/TFA/TEA (100/0.1/0.1) B) water/TFA/TEA (100/0.1/0.1) 0-50% B (0-4 min), 50% B (4-5 min)


Eluent ： A) water/TFA (100/0.1) B) acetonitrile/TFA (100/0.1) 0-30% B (0-4 min), 30% B (4-5 min)


1. TRP-GLY (MW 261)2. GLY-GLY-PHE (MW 279)3. Angiotensin I (MW 1,296)4. Substance P (MW 1,348) 5. Insulin (bovine) (MW 5,700)

2,1

3

4

5

0 2 4 6 8 min

A991127I

YMC-Pack ODS-A（5 µm, 120 Å）50×4.6 mmI.D.

※Shipping solvent for Diol-NP is hexane/2-propanol (99.5/0.5). In case of using mobile phase including water, take care of miscibility.※For details and ordering information, see P86, 87.

Peptides

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Normal-phase columns

YMC-Pack Polyamine Ⅱ

A m i n o c o l u m n w i t h i m p r o v e d d u r a b i l i t yYMC-Pack Polyamine Ⅱ is a silica-based packing bonded with polyamine. It is particularly useful for separation of sugars. The column lifetime of YMC-Pack Polyamine Ⅱ in aqueous mobile phase is longer than conventional silica-based amino columns, and thus is

applicable to separation of oligosaccharides using mobile phase with relatively higher water content. In addition, YMC-Pack Polyamine Ⅱ can be used to separate ionic compounds with a combination of normal-phase mode and weak anion exchange mode.

■Pore size：120 Å■Usable pH range：2.0 〜 7.5

● Silica gel chemically bonded with polyamine● The most suitable column for separation of sugars● Solves durability problem of conventional

silica-based amino columns

● Useful for separation of hydrophilic compounds including vitamins

● Useful for separation of fat-soluble compounds using nonaqueous mobile phase

Suitable column for separation of sugars

1

23

10 20 300 minF030618H

4

5

Column ： YMC-Pack Polyamine Ⅱ 250×4.6 mmI.D.

Eluent ：acetonitrile/water (75/25)Flow rate ：1.0 mL/minTemperature ：25 ℃Detection ：RI, 32×10-6 RIU/FS

※For details and ordering information, see P88, 89.

Elution volume of sugars and sugar alcohols

Arabitol

65

60

55

50

45

40

35

30

25

20

15

10

5

080%75% 85%

Acetonitrile concentration (%)

※Not detected with 85% acetonitrile

Elu

tion

volu

me

(mL)

Inositol

Palatinose

SucraloseSucralose

FucoseFucose

Fructose

Mannitol

Glucose

Galactose（※）

Mannose（※）

Sucrose

Maltitol

Raffinose

Maltose

Sucrose

Maltitol

Raffinose

Maltose

LactoseLactose

Palatinit

Lactitol

Sorbitol

Mannitol

Glucose

Sorbitol

XyloseXylose

Xylitol

GlycerolGlycerol

RhamnoseRhamnose

ErythritolErythritol

Ribitol

Column ： YMC-Pack Polyamine Ⅱ 250×4.6 mmI.D.

Temperature ：25 ℃

Excellent durability

120

100

80

60

40

20

00 500 1000 1500 2000 2500 3000 3500 4000

Polyamine Ⅱ

Column volumes

Silica based amino column

Rat

e of

initi

al r

eten

tion

of L

acto

se (%

)

YMC-Pack Polyamine Ⅱ shows little change in retention time of lactose after passing 3,000 column volumes of 75% acetonitrile mobile phase. On the other hand, retention time on conventional silica-based amino column decreases to 80% of the initial value continuously from the beginning.

Durability in 75% acetonitrile mobile phase condition

1. Fructose2. Glucose3. Sucrose4. Maltose5. Lactose

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columns and bulk packings for bio-macro molecular ... general catalog 5 columns and bulk packings...

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