HPLC Columns SHOWA DENKO K.K. Capture the Essence.

Transcription

1 HPLC Columns 0 0 Capture the Essence SHOWA DENKO K.K. US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

2 TM We provide a wide range of products to meet your analytical needs, from pretreatment and separation columns to calibration standards for size exclusion chromatography. Please visit the Shodex website to see application data and uses. The following names are trademarks or registered trademarks of SHOWA DENKO K.K. Shodex, AFpak, Asahipak, AXpak, CLNpak, CXpak, HILICpak, MSpak, ODP, OHpak, ORpak, RSpak, SUGAR, USPpak [Caution]. Please read the operating manual included on the product carefully before use.. For improvement purposes, some specifications are subject to change without notice.. Provided to help you select the appropriate column, the figures and descriptions in this catalogue are not guaranteed and do not warrant suitability for your applications.. It is essential to take normal precautions when handling reagents and other chemical products even if the safety information is not included on the operating manual.. Products described in this brochure are not intended for medical use or medical applications including medical diagnosis. US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

3 Contents Column selection Analysis and preparative columns Reversed Phase, Hydrophilic Interaction and Normal Phase Chromatography Ligand Exchange Chromatography Ion Exclusion Chromatography Sample pretreatment columns Notice of change of product name Information Index Ion Chromatography Size Exclusion Chromatography Calibration Standards for SEC Ion Exchange Chromatography Columns for Special Separation Modes Types of Columns, Base Materials, Functional Groups and Ligands HPLC Separation Modes Column Selection by Sample Character and Separation Mode Column Selection (Application) Comparison of the Features Among Shodex Reverse Phase Chromatography (RPC) Columns Columns for Polymer-based Reversed Phase Chromatography (ODP HP) Columns for Polymer-based Reversed Phase Chromatography (Asahipak) 0 Columns for Polymer-based Reversed Phase Chromatography (Rspak) Columns for Silica-based Reversed Phase Chromatography (ODS Columns) Columns for Silica-based Reversed Phase Chromatography (Other Columns) Columns for Silica-based HILIC and Normal Phase Chromatography Columns for Polymer-based Hydrophilic Interaction Chromatography (HILIC) (HILICpak) 0 Columns for Polymer-based Hydrophilic Interaction Chromatography (HILIC) (Asahipak) Columns for Ligand Exchange Chromatography Columns for Ion Exclusion Chromatography Columns for Ion Chromatography (Anion Analysis) 0 Columns for Ion Chromatography (Cation Analysis) Column Selection for Size Exclusion Chromatography (SEC) Precautions for Polar Polymer Analysis Aqueous SEC (GFC) Columns : Silica-based Aqueous SEC (GFC) Columns : Polymer-based Multimode Columns Aqueous/Organic SEC Columns Organic SEC (GPC) Columns (General Analysis) : THF Organic SEC (GPC) Columns (General Analysis) : Chloroform Organic SEC (GPC) Columns (General Analysis) : DMF 0 Solvent-peak Separation Columns for Organic SEC (GPC) 0 Organic SEC (GPC) Columns : Rapid Analysis Organic SEC (GPC) Columns : High Performance Analysis Organic SEC (GPC) Columns : For Preparative Organic SEC (GPC) Columns : Linear Calibration Type Organic SEC (GPC) Columns : High Temperature/Ultra High Temperature Analysis Organic SEC (GPC) Columns : HFIP 0 Applicability of SEC (GPC) Columns to Solvent Replacement Calibration Standards for SEC Columns for Anion Exchange Chromatography Columns for Cation Exchange Chromatography Column for Hydrophobic Interaction Chromatography Columns for Affinity Chromatography Columns for Chiral Separation Column for High Temperature Reversed Phase Chromatography Columns for GPC Clean-up 0 Pretreatment Columns for Column Switching Method Notice of Changes USP (Ver.) Column List Column Cleaning Procedures General Precautions for Column Handling Column Trouble Shooting HPLC System Trouble Shooting 9 Index by 0 Index by US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

4 Types of Columns, Base Materials, Functional Groups and Ligands Separation Type Base Material Reversed Phase & HILIC (Polymer-based) Reversed Phase & HILIC & Normal Phase (Silica-based) Ligand Exchange Ion Exclusion Ion Chromatography Aqueous SEC (GFC) Multimode Aqueous/Organic SEC Organic SEC (GPC) Ion Exchange Hydrophobic Interaction Affinity Chiral Separation GPC Clean-up Column Switching Pretreatment Asahipak Asahipak Asahipak RSpak RSpak RSpak RSpak RSpak HILICpak HILICpak Asahipak Silica Silica Silica Silica Silica Silica Silica Silica SUGAR SUGAR SUGAR RSpak SUGAR EP USPpak SUGAR RSpak IC IC IC IC IC PROTEIN OHpak OHpak Asahipak Asahipak MSpak IEC IEC IEC PIKESS Asahipak AXpak IEC IEC PIKESS IEC Asahipak CXpak HIC AFpak ORpak ORpak CLNpak CLNpak MSpak MSpak ODP HP ODP-0, ODP-0 CP-0 CP-0 RP-, DS DE DM- NN JJ-0 VG-0 VT-0 NHP ET-RP C CM, CP C C CN NPE PYE SIL NH SC SP00 KS-00 DC- SZ SC0-F MN- SH KC- NI-, I-A SI YS-0 YK- Y-, T- KW-00 KW00 SB-00 HQ LB-00 GS-HQ GF-HQ GF-0 KF-00, K-00, KD-00, KF-00, KF-00HQ, LF, HT-00, UT-00, AT-0MSHFIP-00, HFIP-00 QA- DEAE- DEAEN DEAE-B ES-0N WA- SP- SP-0N SP-B CM- ES-0C P-S PH- Various CDBS- CRX- EV PAE PK GF-A Polyhydroxymethacrylate Polyvinyl alcohol Polyvinyl alcohol Polyvinyl alcohol Styrene divinylbenzene copolymer Polymethacrylate Polyhydroxymethacrylate Polyhydroxymethacrylate Polyvinyl alcohol Polyvinyl alcohol Polyvinyl alcohol Polyvinyl alcohol Polyvinyl alcohol Silica Silica Silica Silica Silica Silica Silica Silica Silica Styrene divinylbenzene copolymer Styrene divinylbenzene copolymer Styrene divinylbenzene copolymer Styrene divinylbenzene copolymer Styrene divinylbenzene copolymer Styrene divinylbenzene copolymer Styrene divinylbenzene copolymer Styrene divinylbenzene copolymer Styrene divinylbenzene copolymer Polyhydroxymethacrylate Polyvinyl alcohol Polyvinyl alcohol Silica Styrene divinylbenzene copolymer Silica Silica Polyhydroxymethacrylate Polyhydroxymethacrylate Polyvinyl alcohol Polyvinyl alcohol Polyvinyl alcohol Styrene divinylbenzene copolymer Polyhydroxymethacrylate Polyhydroxymethacrylate Polyhydroxymethacrylate Polyhydroxymethacrylate Polyvinyl alcohol Polyhydroxymethacrylate Polyhydroxymethacrylate Polyhydroxymethacrylate Polyhydroxymethacrylate Polyhydroxymethacrylate Polyvinyl alcohol Styrene divinylbenzene copolymer Polyhydroxymethacrylate Polyhydroxymethacrylate Silica Polyhydroxymethacrylate Styrene divinylbenzene copolymer Polyvinyl alcohol Hydrophilic copolymers containing N-vinyl acetamide Polyvinyl alcohol Functional Group, Ligand Page Octadecyl Octyl Butyl Sulfo Quaternary ammonium Ao Quaternary ammonium Ao Octadecyl Octadecyl Octadecyl Octyl Butyl Cyanopropyl Nitrophenylethyl Pyrenylethyl Aopropyl Sulfo(Ca + ) Sulfo(Pb + ) Sulfo(Na + ) Sulfo(Na + ) Sulfo(Zn + ) Sulfo(Ca + ) Sulfo(Ca + ) Sulfo Sulfo Quaternary ammonium 0 Quaternary ammonium 0 Carboxyl Carboxyl Sulfo Hydrophilic polymer Hydrophilic polymer,, 0,,,, 0 Quaternary ammonium Diethylaoethyl Diethylaoethyl Diethylaoethyl Diethylaoethyl Diethylaoethyl Sulfopropyl Sulfopropyl Sulfopropyl Carboxymethyl Carboxymethyl Sulfo(Na + ) Phenyl Various ligand β-cyclodextrin derivative L-Ao acid derivative 0 0 US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

5 HPLC Separation Modes Liquid chromatography (LC) uses liquid as mobile phase (eluent). It is an analytical method that separates a mixture of compounds based on their physical and chemical differences. High performance liquid chromatography (HPLC) is a method that introduces the mobile phase under high-pressure conditions resulting in rapid and high-performance separations. The various interactions between the analyte, stationary phase (packing material), and mobile phase are the key factors for the separation. A wide variety of separation modes can be achieved by using particular combinations of stationary and mobile phases. Separation mode Reversed Phase Chromatography (RPC) Hydrophilic Interaction Chromatography (HILIC) Normal Phase Chromatography (NPC) Ligand Exchange Chromatography (LEX) Ion Exclusion Chromatography (IEX) Ion Chromatography (IC) Size Exclusion Chromatography (SEC) Ion Exchange Chromatography (IEC) Hydrophobic Interaction Chromatography (HIC) Affinity Chromatography (AFC) Chiral Separation Chromatography (CS) Multimode Chromatography Separation is based on hydrophilic interaction. A high polarity stationary phase is used. Typically the mobile phase contains a mixture of organic solvents such as acetonitrile and aqueous solvents (water or buffer). Using the higher polarity mobile phase causes a faster elution. Applicable for the analysis of high polar substances. Separation is based on the partition equilibrium between the stationary phase and the mobile phase. The polarity of the stationary phase is higher than that of the mobile phase. Typically the mobile phase contains a mixture of organic solvents with different polarities such as hexane and isopropanol. Using the higher polarity mobile phase causes a faster elution. Separation is based on differences in analytes coordination complex. Stationary phase modified with metal sulfonate complex ion. Works in combination with size exclusion or HILIC modes. Separation is based on electrostatic interaction (repulsion) between the ion exchanger and ionic solutes. Dissociated ionic molecules elute faster than non-dissociated forms. Used mainly for the analysis of organic acids. Separation is based on electrostatic interaction (bonding) between the ion exchanger and ionic solutes. Has a relatively small ion exchange capacity. Electrical conductivity detector can be used with low-salt concentration mobile phase. Used mainly for the analysis of inorganic compounds. Network or pores on the surface of the packing material works as molecular sieve to separate molecules based on their sizes. To separate molecules solely based on their sizes, it requires an analytical condition without any analyte and packing gel interaction. The bigger the molecule size, the faster the elution sequence. Used for molecular weight or molecular distribution deteration of macromolecules and qualification of oligomers. Separation is based on electrostatic interactions between the ion exchanger and ionic solutes. The mobile phase of choice should have a sufficient buffering capacity at the ph that produces the largest charge differences between the analyte of interest. The elution position is optimized by varying the ph, salt concentration, and/or ionic strength of the mobile phase. Separation is based on hydrophobic interaction. Hydrophobic functional group is modified on the stationary phase. Adsorption of analytes generally occurs at a high salt concentration and they are released by lowering the salt concentration. Used mainly for the analysis of proteins. Separation is based on adsorption of the analyte to the specific biologically derived ligand pair. Highly selective. A buffer solution with the appropriate ph and ionic strength is selected based on the type of ligand, analytes, and their interaction. Used mainly for the purification and concentration of biological active substances. Separation of optical isomers using chiral selectors. Highly selective. Separation is based on the combination of different modes. Column Selection by Sample Character and Separation Mode Sample Solubility Aqueous soluble Sample MW,000,000 Separation Mode RPC LEX IEX SEC IEC HIC AFC RPC HILIC LEX IEX IC SEC IEC AFC CS Sample Solubility Organic soluble Sample MW,000,000 Characteristics Separation is based on the partition equilibrium between stationary phase and mobile phase. The polarity of the stationary phase is lower than that of the mobile phase. Typically the mobile phase contains a mixture of organic solvents (methanol, acetonitrile, or THF) and aqueous solvents (water or buffer). Using the lower polarity mobile phase causes a faster elution. Separation Mode SEC RPC NPC SEC RPC : Reversed Phase Chromatography HILIC : Hydrophilic Interaction Chromatography NPC : Normal Phase Chromatography LEX : Ligand Exchange Chromatography IEX : Ion Exclusion Chromatography IC : Ion Chromatography SEC : Size Exclusion Chromatography IEC : Ion Exchange Chromatography HIC : Hydrophobic Interaction Chromatography AFC : Affinity Chromatography CS : Chiral Separation Chromatography Types of Columns, Base Materials, Functional Groups and Ligands HPLC Separation Modes Column Selection by Sample Character and Separation Mode US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

6 Column Selection (Application) Pharmaceuticals, Cosmetics Foods Separation Mode Page Separation Mode Page Hydrophobic substances RPC HILIC, 0,,, 0, Monosaccharides Disaccharides Sugar alcohols HILIC LEX+SEC LEX+HILIC 0, Pharmaceuticals Metabolites Additives Hydrophilic substances Substances in bio-fluid (serum plasma urine) Polymer IEC+RPC LEX+SEC RPC SEC+RPC SEC,,,,, 0, Oligosaccharides Low molecular water-soluble dietary fiber Polysaccharides HILIC LEX+HILIC SEC SEC SEC RPC 0,,,,, RPC Organic acids IEX+RPC Moisturizers Polyalcohols LEX+SEC LEX+HILIC SEC, Nutritional ingredients Water-soluble vitas IC RPC IEC+RPC 0, 0, Protein hydrolysates Mucopolysaccharides RPC SEC SEC 0, Fat-soluble vitas HILIC RPC NPC 0, 0 Emulsifiers Preservatives Surfactants Paraben Dehydroacetic acid SEC+RPC SEC RPC, 0,,, Fatty acids Nucleic acids (umami) SEC RPC SEC IEC+SEC IEC+IEX+RPC,,,,,, Optical active materials CS Ao acids IC IEC Food additives Pesticides RPC HILIC RPC IEC+RPC HILIC 0,, 0, 0 Food safety IC 0 Mycotoxin Pretreatment of residual pesticides RPC SEC GPC ( Clean-up ) 0 Separation Mode (Page and Page ) RPC : Reversed Phase Chromatography HILIC : Hydrophilic Interaction Chromatography NPC : Normal Phase Chromatography LEX : Ligand Exchange Chromatography IEX : Ion Exclusion Chromatography IC : Ion Chromatography SEC : Size Exclusion Chromatography IEC : Ion Exchange Chromatography HIC : Hydrophobic Interaction Chromatography AFC : Affinity Chromatography CS : Chiral Separation Chromatography New Materials Synthetic polymers Additives Oligomers Organic solvent soluble Polar organic solvent soluble High temperature/ Ultra high temperature Water-soluble Organic solvent soluble Polar organic solvent soluble Water-soluble Separation Mode SEC RPC SEC Page,,,,,, 0,,,,, 0,,,,,,,, 0,,,, US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

7 Biotechnology Genomics Nucleobases Nucleotides Nucleosides Oligo nucleic acids DNA/RNA Separation Mode RPC IEC+SEC IEC RPC IEC+SEC IEC SEC Page,, Environment Water quality Anions Oxyhalides Cyanide Cyanogen chloride Cations Surfactants Separation Mode IC IC IEC+HILIC IEX IC RPC SEC+RPC Page , Column Selection (Application) Ao acids RPC IEC+IEX+RPC IEC 0 Perchloric acids IC IEC+HILIC RPC 0 0,, Proteomics IEC+SEC RPC 0, Pesticides IEC+RPC HILIC 0 Peptides Proteins SEC IEC,,,, Anions IC IC 0 0 HIC Heavy metals IC RPC 0, Humic substances SEC Glycoproteins SEC IEC,,,, Soil Organic arsenic IEX+RPC RPC,, HIC AFC Pesticides IEC+RPC HILIC 0 Glycomics Sugar chains Monosaccharides HILIC AFC HILIC LEX+SEC 0, 0, Environmental hormones Pretreatment of Phthalates PCBs Benzo [a] pyrene IC SEC GPC ( Clean-up ) 0 0 Hormones Sialic acids Uronic acids Aldonic acids Aes LEX+HILIC IEX+RPC RPC IEC RPC, 0, 0 Bioethanols Monosaccharides Oligosaccharides Oligosaccharides Alcohols Furfural Saccharides Organic acids Alcohols Furfural HILIC LEX+SEC LEX+SEC IEX+RPC+SEC 0, Steroids HILIC SEC 0,, Hemicelluloses Celluloses SEC 0, Lipids Phospholipids Lipoproteins NPC SEC SEC AFC,, Biodiesels Cations Fatty acid glycerides Fatty acid methyl esters Organic acids IC SEC RPC IC 0 US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

8 Comparison of the Features Among Shodex Reverse Phase Chromatography (RPC) Columns ODS columns are the most popular reversed phase columns that are packed with silica-based octadecyl group. Shodex provides not only ODS columns but also polymer-based reversed phase columns with different functional groups. You can choose a suitable column for your application based on the features listed below. Features ODP HP ODP-0 CP-0 CP-0 ODP-0 RP- DS- DS- DE DM- NN JJ-0 C CM CP C C CN NPE PYE Provides a large theoretical plate number nearly twice as much as generally available polymer-based reversed phase columns Offers enhanced retention of high polar substances compared to ODS columns Suitable for the analysis of small molecules such as pharmaceuticals in the presence of protein matrix Ideal for LC/MS analysis of high polar compounds Corresponds to USP L9 Relatively large pore size is suitable for the analysis of ao acids, peptides, and proteins Usable in a wide ph range from ph to Usable in 00% water or buffer solution Best used for the analysis of basic substances ODP-0 corresponds to USP L Higher performance type of ODP-0 series Corresponds to USP L Large pore size is suitable for the analysis of proteins and peptides Corresponds to USP L Suitable for reversed phase analysis of highly hydrophilic substances, that are not well retained by ODS columns Corresponds to USP L Polymer-based columns, with similar polarity to that of ODS columns, can be used in general purposes Wide working ph range (form ph to ), usable in 00% water and buffer solutions Corresponds to USP L Suitable for the analysis of ao acids and water-soluble vitas Corresponds to USP L9 The packing material contains sulfo groups, and supports multimode (reversed phase and cation exchange) analysis Ideal for the analysis of complex samples containing neutral and ionic substances The packing material contains trace amounts of quaternary ammonium groups, and supports multimode (reversed phase and anion exchange) analysis Ideal for analysis of complex samples containing neutral and ionic substances Fully end capped ODS column available at very reasonable price Corresponds to USP L Monomeric type ODS column, fully end capped high purity silica (99.99% or higher) Corresponds to USP L Polymeric type ODS column, fully end capped high purity silica (99.99% or higher) Excellent acid tolerance Advantageous for separating planar and nonplanar compounds from each other Corresponds to USP L Use when the retention capacity of C is too strong or that of C is too week Applicable to ion pair chromatography, because of its rapid mass transfer and equilibrium Corresponds to USP L Use when the retention capacity of C or C is too strong Corresponds to USP L Utilizes reversed phase interaction and π-electron interaction to separate regioisomers, which typically cannot be separated with ODS, C, or C columns Corresponds to USP L0 Utilizes several types of interactions based on π-electrons to separate structural isomers NPE corresponds to USP L US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

9 The interrelation between hydrophobicity and retention, and the interrelation between steric selectivity and retention were compared among Shodex columns for reversed phase chromatography. The retention factor (k ) of amylbenzene was used as the retention, the separation factor (α) between n-butylbenzene and n-ao benzene was used as the hydrophobicity. The separation factor between o-terphenil and triphenylene was used as the steric recognition. Lager separation factor means higher hydrophobicity and higher steric selectivity. Interrelation between hydrophobicity and retention Hydrophobicity EG C C C C CM CP PYE C C ODP-0 NPE CN CP-0 DE- ODP HP C0 C C CP-0 Retention Column size :.mm I.D. x 0mm each Eluent : H O/CH OH=0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C Comparison of separation of alkylalcohol due to different functional groups Steric selectivity ODP HP CN NPE CP-0 C n-buthylbenzene o-terphenyl PYE DE- CP-0 C Retention Column size :.mm I.D. x 0mm each Eluent : H O/CH OH=0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C ODP-0 CP C n-amylbenzene Triphenylene Interrelation between steric selectivity and retention CM Comparison of separation due to the difference in steric selectivity Sample : Ethyleneglycol, n-alkylalcohol Sample : µl. n-buthylbenzene. n-amylbenzene CP-0 (Butyl). o-terphenyl CM D. Triphenylene ODP-0 Comparison of the Features Among Shodex Reverse Phase Chromatography (RPC) Columns EG C C C CP-0 (Octyl) C0 C C Elution volume (ml) 0 0 Carbon number (n) CP-0 CP-0 Relationship between the carbon number of alkylalcohols and elution volume PYE D EG C C 0 C ODP-0 (Octadecyl) C0 C C 0 0 Column : Shodex Asahipak ODP-0 D Shodex Asahipak CP-0 D Shodex Asahipak CP-0 D Eluent : H O/CH OH=0/0 Flow rate : 0.mL/ Column temp. : 0 C Column : Shodex Silica CM D Shodex Silica PYE D Eluent : H O/CH OH=0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

10 Columns for Polymer-based Reversed Phase Chromatography (ODP HP) Please refer to Comparison of the Features Among Shodex Reverse Phase Chromatography (RPC) Columns on page and for features. Standard columns Functional Group Shipping Solvent F00 ODP HP-B, H O/CH CN=/ F00 ODP HP-D, H O/CH CN=/ F00 ODP HP-E, H O/CH CN=/ F00 ODP HPG-A. 0 H O/CH CN=/ F00 ODP HP-B, H O/CH CN=/ F00 ODP HP-D, H O/CH CN=/ F0 ODP HPG-A.0 0 H O/CH CN=/ Base Material : Polyhydroxymethacrylate mm I.D columns [Customized columns] Functional Group F00 ODP HP-B F00 ODP HP-D F0 ODP HPG-A.0 0 Base Material : Polyhydroxymethacrylate Tolerance of ODP HP for alkaline condition Comparison between the chromatograms obtained before and after passing alkaline eluent Before After 00 hr ODP HP-D (.mm I.D.x0mm) Sample : µl. Pyridine 00µg/mL. Phenol 0µg/mL 0 0 Before After hr ODS (.mm I.D.x0mm) 0 0 Correlation between alkaline eluent passing time and relative theoretical plate number Relative theoretical plate number for pyridine (%) ODP HP-D ODS Alkaline eluent passing time (hr) Analysis condition Column : Shodex ODP HP-D ODS from other manufacturer Eluent : H O/CH OH=0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C Eluent passing conditions for alkali tolerance test Column : Shodex ODP HP-D ODS from other manufacturer Eluent : 0mM Sodium phosphate buffer (ph) /CH CN=/ Flow rate : 0.mL/ Column temp. : 0 C US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

11 Comparison between ODP HP and ODP-0 (existing) Sample : µl. Phenol 00mg/L. Methyl benzoate 0mg/L. Toluene 000mg/L. Naphthalene 0mg/L ODP HP-D N:,00 ODP-0 D (existing) Column : Shodex ODP HP-D Eluent : H O/CH CN=/ Flow rate : 0.mL/ Detector : UV (nm) Column temp. : 0 C Ratio of Pressure (%) * ODS (.0mm I.D. 0mm) ODP HP-B (.0mm I.D. 0mm) Number of Injection * Considering original pressure as 00% Column : Shodex ODP HP-B ODS from other manufacturer Eluent : mm Ammonium acetate aq./ch CN=90/0 Flow rate : 0.mL/ Detector : UV (0nm) Column temp. : 0 C N:,00 Column : Shodex Asahipak ODP-0 D Eluent : H O/CH CN=/ Flow rate : 0.mL/ Detector : UV (nm) Column temp. : 0 C Influence of repeated protein injection on column pressure ODP HP columns are packed with gels with increased surface polarity and smaller pore size which prevent the adsorption of proteins. BSA was injected multiple times to both ODS and ODP HP columns. A significant column pressure increase was observed for the ODS column, while no considerable change was observed for the ODP HP column even after 0 injections. Sample : µl BSA.0mg/mL Imidazoles. Sample : 0.% each, 0µL. Imidazole. -Methylimidazole. -Methylimidazole Column : Shodex ODP HP-E Eluent : 0mM Na HPO aq./ch CN=90/0 Flow rate : 0.mL/ Detector : UV (0nm) Column temp. : 0 C Anticonvulsant in serum Sample : 0µL. -Acetaophenol (I.S.) 0µg/mL. Zonisamide.0µg/mL. Phenobarbital 9.0µg/mL. Carbamazepine.µg/mL. Phenytoin 9.0µg/mL Column : Shodex ODP HP-E Eluent : mm Sodium phosphate buffer (ph.)/ch CN=0/0 Flow rate : 0.mL/ Detector : UV (0nm) Column temp. : 0 C Sample pretreatment : Mix the same volumes of serum and acetonitrile. Centrifuge the mixture at 000g for utes. Use the supernatant as sample. Data courtesy of Katsuko Hara.MT Yutaka Komiyama Ph.D., Department of Clinical Sciences and Laboratory Medicine, Kansai Medical University. Columns for Polymer-based Reversed Phase Chromatography (ODP HP) Barbital recovery rate comparison of ODP HP-B and ODS in the presence of BSA ODP HP Barbital 00ng/mL ODS Barbital 00ng/mL For the analysis of drugs in samples containing proteins in matrix using LC/MS, ODP HP columns show less matrix effects (ion suppression in this case) compared to ODS columns due to fact that proteins can not be retained on ODP HP columns and are eluted as a void. Barbital Barbital Peak Area : 00% Peak Area : 00% Barbital 00ng/mL in BSA.0mg/mL Barbital 00ng/mL in BSA.0mg/mL Barbital Barbital Peak Area : 99% Peak Area : % Column : Shodex ODP HP-B ODS from other manufacturer Eluent : 0mM Ammonium acetate aq. /CH CN=0/0 Flow rate : 0.mL/ Detector : ESI-MS (SIM Negative : m/z ) Column temp. : 0 C Injection vol. : 0µL 9 US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

12 Columns for Polymer-based Reversed Phase Chromatography (Asahipak) Please refer to Comparison of the Features Among Shodex Reverse Phase Chromatography (RPC) Columns on page and for features. Standard columns Functional Group Column Size (mm) Shipping Solvent F00 Asahipak ODP-0 D,000 Octadecyl 0. 0 H O/CH CN=/ F00 Asahipak ODP-0 E,000 Octadecyl 0. 0 H O/CH CN=/ F000 Asahipak ODP-0 D 9,000 Octadecyl H O/CH CN=/ F000 Asahipak ODP-0 E,000 Octadecyl H O/CH CN=/ F000 Asahipak ODP-0G A Octadecyl.0 0 H O/CH CN=/ F00 Asahipak ODP-0 B,00 Octadecyl 0. 0 H O/CH CN=/ F000 Asahipak ODP-0 D 9,000 Octadecyl 0. 0 H O/CH CN=/ F000 Asahipak ODP-0 E,000 Octadecyl 0. 0 H O/CH CN=/ F00 Asahipak ODP-0G A Octadecyl. 0 H O/CH CN=/ F0009 Asahipak ODP-0 D,000 Octadecyl H O/CH CN=/ F00 Asahipak ODP-0G A Octadecyl.0 0 H O/CH CN=/ F000 Asahipak CP-0 D,000 Octyl 0. 0 H O/CH CN=/ F000 Asahipak CP-0 E,000 Octyl 0. 0 H O/CH CN=/ F000 Asahipak CP-0G A Octyl. 0 H O/CH CN=/ F000 Asahipak CP-0 D,000 Butyl 0. 0 H O/CH CN=/ F000 Asahipak CP-0 E 9,000 Butyl 0. 0 H O/CH CN=/ F000 Asahipak CP-0G A Butyl. 0 H O/CH CN=/ mm I.D columns [Customized columns] Base Material : Polyvinyl alcohol Functional Group F0 Asahipak ODP-0 B Octadecyl F0 Asahipak ODP-0 D Octadecyl F0 Asahipak ODP-0G A Octadecyl Semi-micro columns * The following semi-micro columns are made to order. Functional Group F0 ODP0-B Octadecyl F0 ODP0-D Octadecyl Preparative columns * Preparative columns are made to order. Standard Column F000 Asahipak ODP-0 0E 0, ODP-0, ODP-0 F00 Asahipak ODP-90 0F 9, ODP-0, ODP-0 F000 Asahipak ODP-0G B. 0 F000 Asahipak CP-0 0E, CP-0 F00 Asahipak CP-0G B. 0 0 F000 F00 Asahipak CP-0 0E Asahipak CP-0G B, CP-0 US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

13 Tolerance of ODP-0 for alkaline condition Scopolae m/z 0(+) 0hr 90hr 0hr 0hr Column : Shodex Asahipak ODP-0 D Eluent : 0mM NaOH aq. (ph.0)/ch CN=/ Flow rate : 0.mL/ Detector : UV (nm) Column temp. : 0 C HC N O O O OH Sample :. Acetophenone. Butyrophenone. Hexanophenone. Heptanophenone. Octanophenone LC/MS analysis of basic drugs Sample : 0ng/mL each, µl. Scopolae. Atropine Local anesthetics Dissociation of tertiary ao groups in basic drugs can be suppressed by making ph of the eluent higher than pka of the ao groups. This increases the relative hydrophobicity of the basic drugs, thereby allowing the column to retain the drugs stronger and provide baseline separation of them. ph 0 0 Fat-soluble vitas ph 0 0 Column : Shodex AsahipakODP-0 D Eluent : mm Phosphate buffer/ch CN=0/0 Flow rate : 0.mL/ Detector : UV (nm) Column temp. : 0 C ph 0 0 Sample :. Benzocaine HN. Lidocaine CH CH. Tetracaine NHCOCHN COOCHCHN NH(CH)CH COOCH Sample : 0µL. Vita K.µg/mL. Vita A.0 IU/mL. Vita A acetate 0. IU/mL. Vita D.µg/mL. Vita D. IU/mL. Vita E acetate.µg/ml. Vita E.µg/mL. Vita K.µg/mL CH CH CH CH Columns for Polymer-based Reversed Phase Chromatography (Asahipak) Atropine m/z 90(+) N O O OH Column : Shodex ODP0-D Eluent : 0.0% Ammonia aq./ch CN=0/0 Flow rate : 0.mL/ Detector : ESI-MS (SIM Positive) Column temp. : 0 C Column : Shodex Asahipak ODP-0 E Eluent : CH CN/CH OH=0/0 Flow rate : 0.mL/ Detector : UV (0nm) Column temp. : 0 C Analysis of azithromycin according to JP method Macrolide antibiotics Gradient analysis of proteins and peptides Sample : µl. Azithromycin mg/ml.,'-bis(diethylao)benzophenone 0µg/mL Resolution (Azithromycin/Internal standard) Sample : 0.% each, 0µL. Erythromycin. Azithromycin 0%CHCN 9 0 Sample : 0µL No %CHCN Sample Lys-Bradykinin Bradykinin Met-Enkephalin Neurotensin Leu-Enkephalin Substance P Bacitracin Insulin Insulin B chain Lysozyme Mastoparan Myoglobin Recovery MW (%) Column : Shodex Asahipak ODP-0 E Eluent : 0mM K HPO aq. (ph.0 adjusted with potassium hydroxide aq.)/ch CN=0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C Column : Shodex Asahipak ODP-0 E Eluent : 0mM Potassium phosphate buffer (ph.0)/ch CN=0/0 Flow rate : 0.mL/ Detector : UV (nm) Column temp. : 0 C Column : Shodex Asahipak ODP-0 D Eluent : (A); 0.0% TFA aq./ch CN=0/0 (B); 0.0% TFA aq./ch CN=0/0 Linear gradient; (A) to (B), 0 Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

14 Columns for Polymer-based Reversed Phase Chromatography (RSpak) Please refer to Comparison of the Features Among Shodex Reverse Phase Chromatography (RPC) Columns on page and for features. Standard columns Functional Group Base Material Particle Size Pore Size Column Size (mm) Shipping Solvent F RSpak RP-,000 Styrene divinylbenzene copolymer 0. 0 H O/CH CN=/9 F09 RSpak RP-G Styrene divinylbenzene copolymer. 0 H O/CH CN/THF=0/0/0 F0000 RSpak DS-,00 Styrene divinylbenzene copolymer H O/CH CN/THF=0/0/0 F000 RSpak DS-,000 Styrene divinylbenzene copolymer H O/CH CN/THF=0/0/0 F000 RSpak DS-G Styrene divinylbenzene copolymer 0. 0 H O/CH CN/THF=0/0/0 F0000 RSpak DE-,000 Polymethacrylate.0 0 H O F0000 RSpak DE-,000 Polymethacrylate. 0 H O/CH CN=0/0 F00900 RSpak DE-L,000 Polymethacrylate. 0 H O/CH CN=0/0 F000 RSpak DE-G A (RSpak DE-G) Polymethacrylate 0. 0 H O F0000 RSpak DE-,000 Polymethacrylate.0 0 H O/CH CN=0/0 F00 RSpak DE-G A (RSpak DE-SG) Polymethacrylate.0 0 H O/CH CN=0/0 F0000 RSpak DM-,00 Polyhydroxymethacrylate mm H PO aq. F000 RSpak DM-G A (RSpak DM-G) Polyhydroxymethacrylate. 0 mm H PO aq. F000 RSpak NN- 9,000 Sulfo Polyhydroxymethacrylate M Sodium phosphate buffer (ph.0) F000 RSpak NN-,000 Sulfo Polyhydroxymethacrylate M Sodium phosphate buffer (ph.0) F000 RSpak NN-G Sulfo Polyhydroxymethacrylate M Sodium phosphate buffer (ph.0) F000 RSpak NN-,000 Sulfo Polyhydroxymethacrylate M Sodium phosphate buffer (ph.0) F000 RSpak JJ-0 D,00 Quaternary ammonium Polyvinyl alcohol H O/CH CN=0/0 F000 RSpak JJ-0 D,00 Quaternary ammonium Polyvinyl alcohol H O/CH CN=0/0 US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

15 Semi-micro columns * The following semi-micro columns are made to order. F0 F0 F0 DE-B DE-E NN-D Functional Group Sulfo 0,000 Base Material Polymethacrylate Polymethacrylate Polyhydroxymethacrylate Preparative columns * Preparative columns are made to order. F0 F0090 F0 F009 RSpak DE-0 RSpak DE-G B (RSpak DE-LG) RSpak DE-0 RSpak DE-G 0C (RSpak DE-LLG) Standard Column DE-, DE- DE-, DE- DE-, DE- Columns for Polymer-based Reversed Phase Chromatography (RSpak) F0 RSpak DM-0, DM- F000 RSpak DM-G B (RSpak DM-LG).0 0 F0 RSpak DM DM- F00 RSpak DM-G 0C (RSpak DM-LLG) US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

16 Separation and recovery rate of standard proteins Alkylbenzenes Recovery (%). Ribonuclease A 9. Insulin 9. Cytochrome c 00. Lysozyme 00. BSA 9. Myoglobin 0. Ovalbu Sample : µl. m-cresol 0.%.,-Xylenol 0.%. Benzene 0.%. Toluene 0.%. Ethylbenzene 0.%. n-propylbenzene 0.% Column : Shodex RSpak RP- Eluent : (A); 0.% TFA aq./ch CN=99/ (B); 0.% TFA aq./ch CN=/9 Linear gradient; (B%) 0% to 0%, 0 Flow rate :.0mL/ Detector : UV (0nm) Column temp. : Room temp. 0 0 Column : Shodex RSpak DS- Eluent : H O/CH CN/THF=0/0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C Fatty acid methyl esters Organic acids Sample : 0.% each, 0µL. Methyl linoleate. Methyl palmitate. Methyl oleate. Methyl stearate Sample : µl. Glyoxylic acid.mg/ml. Tartaric acid.9mg/ml. Malic acid.0mg/ml. Lactic acid µl/ml. Malonic acid.9mg/ml. Acetic acid µl/ml. Succinic acid.0mg/ml. Levulinic acid.9mg/ml 9. Propionic acid µl/ml Column : Shodex RSpak DS- Eluent : H O/CH CN/THF=//0 Flow rate :.0mL/ Detetor : RI Column temp. : 0 C Column : Shodex RSpak DE- Eluent : 0mM H PO aq. Flow rate :.0mL/ Column temp. : 0 C Food additives (Preservatives) Diols Sample : 0µL. Saccharin sodium 0.00%. p-hydroxybenzoic acid 0.00%. Sorbic acid 0.0%. Benzoic acid 0.0%. Methyl paraben 0.0%. Dehydroacetic acid 0.0%. Ethyl paraben 0.0%. Propyl paraben 0.0% n= Sample : % each,.µl HO(CH)nOH Column : Shodex RSpak DE- Eluent : 0mM KH PO + 0.% H PO aq./ch CN=/ Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C 0 0 Column : Shodex RSpak DE- Eluent : H O Flow rate :.0mL/ Column temp. : 0 C US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

17 LC/MS analysis of organic acids 0 9 Glyoxylic acid Pyruvic acid Glycolic acid Lactic acid Maleic acid Succinic acid Fumaric acid Adipic acid Sample : 0ng/mL each, 0µL m/z m/z m/z m/z 9 m/z m/z m/z m/z Column : Shodex RSpak DE- Eluent : (A); 0.% (v/v) Formic acid aq. (B); CH CN Linear gradient; B%; % (0) % () % (.) % (0) Flow rate : 0.mL/ Detector : ESI-MS (SIM Negative) Column temp. : 0 C Ao acids Sample : 0.% each, 0µL. Aspartic acid. Glycine. Alanine. Valine. Methionine. Isoleucine Speciation of arsenic Standard arsenic compounds Carnitine Sample : 0µL. L-Carnitine 0.% N + OH O OH Column : Shodex RSpak NN- Eluent : 0.M H PO aq. Flow rate :.0mL/ Detector : UV (0nm) Column temp. : C Sample : Arsenic compounds, 0µL. Monomethylarsonic acid. Arsinic acid. Dimethylarsinic acid. Arsenobetaine. Tetramethylarsonium. Trimethylarsine oxide Columns for Polymer-based Reversed Phase Chromatography (RSpak) sec 000 Urine from an arsenic poisoning patient 000 Urine of a healthy person Column : Shodex RSpak NN- Eluent : 0mM H PO aq. Flow rate :.0mL/ Column temp. : 0 C sec Column : Shodex RSpak NN- Eluent : mm HNO /mm NH NO aq. Flow rate : 0.mL/ Detector : ICP-MS (SIM m/z ) sec Source: Noriko Tsunoda, Pharmacia. 99, vol., No., p.- Phenylenediae isomers LC/MS analysis of haloacetic acids Sample : 00mg/L each, 0µL. p-phenylenediae. m-phenylenediae. o-phenylenediae Standard Sample : 0µL each (Standard) ng/ml each (Tap water). Chloroacetic acid. Dichloroacetic acid. Trichloroacetic acid (.µg/l) Tap water (0.µg/L) Column : Shodex RSpak JJ-0 D Eluent : mm Ammonium acetate buffer (ph9.)/ch CN=0/0 Flow rate : 0.mL/ Detector : UV (nm) Column temp. : 0 C Column : Shodex RSpak JJ-0 D Eluent : mm Ammonium acetate aq. (ph9.)/ch CN=0/0 Flow rate : 0.mL/ Detector : Chloroacetic acid : ESI-MS (SIM Negative : m/z 9) Dichloroacetic acid : ESI-MS/MS (MRM Negative : m/z ) Trichloroacetic acid : ESI-MS (SIM Negative : m/z ) Column temp. : 0 C US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

18 Columns for Silica-based Reversed Phase Chromatography (ODS Columns) Please refer to Comparison of the Features Among Shodex Reverse Phase Chromatography (RPC) Columns on page and for features. Standard columns Functional Group Carbon Load (%) Shipping Solvent F00 C-D,000 Octadecyl 0. 0 H O/CH OH=/ F0 C-E,000 Octadecyl 0. 0 H O/CH OH=/ F000 Silica CM D 0,000 Octadecyl H O/CH OH=0/0 F00 Silica CM E,000 Octadecyl H O/CH OH=0/0 F00 Silica CM D 9,000 Octadecyl H O/CH OH=0/0 F00 Silica CP D 0,000 Octadecyl H O/CH OH=0/0 F00 Silica CP E,000 Octadecyl H O/CH OH=0/0 F00 Silica CP D 9,000 Octadecyl H O/CH OH=0/0 Base Material : Silica Preparative columns * Preparative columns are made to order. Standard Column F000 Silica CM 0E, CM F00 Silica CM 0E, CM US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

19 Columns for Silica-based Reversed Phase Chromatography (Other Columns) Please refer to Comparison of the Features Among Shodex Reverse Phase Chromatography (RPC) Columns on page and for features. Standard columns F00 F00 F00 F00 F00 F009 F00 F00 Standard columns F000 F00 F000 F00 Silica C D Silica C E Silica C D Silica C E Silica CN D Silica CN E Silica NPE D Silica PYE D Silica SIL D Silica SIL E Silica NH D Silica NH E 9,000,000 9,000,000,000,000,000,000 9,000,000,000,000 Functional Group Octyl Octyl Butyl Butyl Cyanopropyl Cyanopropyl Nitrophenylethyl Pyrenylethyl Functional Group Aopropyl Aopropyl Preparative columns * Preparative columns are made to order. F00 F00 F00 F00 Silica C 0E Silica C 0E Silica C 0E Silica C 0E,000,000,000,000 Carbon Load (%) 0 0 Carbon Load (%) Columns for Silica-based HILIC and Normal Phase Chromatography Features SIL Uses high purity silica (99.99% or higher) Suitable for normal phase analysis using a nonpolar organic solvent Corresponds to USP L NH Preparative columns * Preparative columns are made to order. F000 F00 Suitable for saccharides analysis by hydrophilic interaction chromatography (HILIC) Corresponds to USP L Silica SIL 0E Silica SIL 0E,000, Shipping Solvent H O/CH OH=/ H O/CH OH=/ H O/CH OH=/ H O/CH OH=/ H O/CH OH=0/0 H O/CH OH=0/0 H O/CH OH=/ H O/CH OH=0/0 Base Material : Silica Standard Column C C C C Shipping Solvent C H /C H OH=9/ C H /C H OH=9/ H O/CH CN=/9 H O/CH CN=/9 Base Material : Silica Standard Column SIL SIL Columns for Silica-based Reversed Phase Chromatography (ODS Columns) (Other Columns) Columns for Silica-based HILIC and Normal Phase Chromatography F000 F00 Silica NH 0E Silica NH 0E,000, NH NH US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

20 Fat-soluble vitas Anticonvulsant Sample : 0µL. Vita A 0µg/mL. Vita D µg/ml. Vita D µg/ml. Vita E 00µg/mL. Tocopherol Acetate 00µg/mL Sample : 0µL. Ethosuximide 00µg/mL. Phenytoin 0µg/mL. Phenobarbital 0µg/mL. Primidon 0µg/mL. Carbamazepine 0µg/mL 0 0 Column : Shodex C-D Eluent : CH CN Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C 0 0 Column : Shodex C-D Eluent : 00mM Phosphate buffer(ph.) /CH OH/CH CN=// Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C Aflatoxins Trichothecene mycotoxins TFA method Sample : µg/l each, 0µL. Derived Aflatoxin G. Derived Aflatoxin B. Aflatoxin G. Aflatoxin B Sample : 0µL. Nivalenol (NIV). Deoxynivalenol (DON) * mg/l each of DON and NIV were added to wheat sample Reference : Shoku-An No. 0- (August, 0, Japan) "Test Methods Related to Total Aflatoxin" in Notice Reference : Shoku-An No. 000 (July, 00, Japan) Test method of deoxynivalenol Column : Shodex Silica CM E Eluent : H O/CH CN/CH OH=0/0/0 Flow rate :.0mL/ Detector : Fluorescence (Ex. : nm, Em. : 0nm) Column temp. : 0 C Column : Shodex Silica CM E Eluent : H O/CH CN/CH OH=90// Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C Gingerol and shogaol Chlorogenic acid Sample : 0.mg/mL each, 0µL. -Gingerol. -Shogaol Sample : 0.mg/mL each, 0µL. Chlorogenic acid. Caffeine. Caffeic acid Column : Shodex Silica CM D Eluent : (A) ; H O (B) ; CH CN Linear gradient : (B%) 0% to 0% () Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C Column : Shodex Silica CM D Eluent : 0mM H PO aq. /CH OH=0/0 Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

21 Catechins Sample : 0µg/mL each, 0µL. Epigallocatechin. Catechin. Epigallocatechingallate. Epicatechin. Epicatechingallate. Catechingallate 0 0 Dinitronaphthalene isomers 0 0 Column : Shodex Silica CP D Eluent : (A) ; 0mM H PO aq. (B) ; CH CN Linear gradient: (B%) 0% (0 to ), 0 to 0% ( to ), 0% ( to 0) Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C Coenzyme Q0 Column : Shodex Silica CP D Eluent : CH OH/C H OH=/ Flow rate :.ml/ Detector : UV (nm) Column temp. : C 0 Sample :. Naphthalene.,-Dimethylnaphthalene.,-Dinitronaphthalene.,-Dinitronaphthalene Estradiols Column : Shodex Silica NPE D Eluent : H O/CH OH=0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C Sample : µl. Coenzyme Q0 mg/ml Column : Shodex Silica CP D Eluent : H O/CH CN=/ Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C Benzylpyridine isomers Column : Shodex Silica PYE D Eluent : 0mM KH PO aq./ch OH=0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C Sample : 0.mg/mL each, 0µL. Estriol. β-estradiol. Estrone Simultaneous analysis of vita E homologs Column : Shodex Silica SIL D Eluent : n-hexane/isopropanol/acetic acid=000// Flow rate :.0mL/ Detector : Fluorescence (Ex. : 9nm, Em. : nm) Column temp. : 0 C Sample :. -Benzylpyridine. -Benzylpyridine. -Benzylpyridine Sample : 0µL. α-tocopherol µg/ml. α-tocotrienol 0µg/mL. β-tocopherol µg/ml. γ-tocopherol µg/ml. γ-tocotrienol 0µg/mL. δ-tocopherol µg/ml. δ-tocotrienol 0µg/mL Columns for Silica-based Reversed Phase Chromatography (ODS Columns) (Other Columns) Columns for Silica-based HILIC and Normal Phase Chromatography 9 US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

22 Columns for Polymer-based Hydrophilic Interaction Chromatography (HILIC) (HILICpak) Features VG-0 Suitable for saccharides analysis by hydrophilic interaction chromatography (HILIC) High recovery ratio of reducing saccharides Polymer-based packing material provides excellent chemical stability and imum deterioration over extended time period Easily regenerated by washing in a alkaline solution Also suitable for evaporative light scattering detector, corona charged aerosol detector, and LC/MS VT-0 Suitable for anionic substances analysis by hydrophilic interaction chromatography (HILIC) Depends on the eluent selected, the column adds ion exchange mode Polymer-based packing material provides excellent chemical stability and imum deterioration over extended time period Suitable for LC/MS Standard columns VG-0 (Housing Material : SUS) Functional Group Shipping Solvent F000 HILICpak VG-0 D,00 Ao H O/CH CN=0/0 F000 HILICpak VG-0 E,00 Ao H O/CH CN=0/0 F00 HILICpak VG-0G A Ao H O/CH CN=0/0 Base Material : Polyvinyl alcohol (Housing Material : PEEK) Functional Group Shipping Solvent F000 HILICpak VG-0 D,00 Ao H O/CH CN=/ F00 HILICpak VG-0G A Ao H O/CH CN=/ Base Material : Polyvinyl alcohol VT-0 (Housing Material : PEEK) Functional Group Shipping Solvent F000 HILICpak VT-0 D,00 Quaternary ammonium mm HCOONH aq. /CH CN=/ F00 HILICpak VT-0G A Quaternary ammonium mm HCOONH aq. /CH CN=/ Base Material : Polyvinyl alcohol 0 US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

23 Recovery of reducing sugar Sample : mg/ml each, µl. Fructose. Mannose. Glucose. Sucrose VG-0 D (.mm I.D. x 0mm) Silica-based ao column (company-a) (.mm I.D. x 0mm) Silica-based ao column (company-b) (.mm I.D. x 0mm) Column : Shodex HILICpak VG-0 D VG-0 D Silica-based Silica-based Ao Column Ao Column Silica based ao columns from other manufacturers (company-a) (company-b) Eluent : H O/CH CN=0/0 Flow rate : 0.mL/ (VG-0 D).0mL/ (Silica based ao column) Column temp. : 0 C Lactose, epilactose, and aactulose When an ao column is used for analyzing saccharides, the recovery ratio of reducing saccharides such as mannose, arabinose or xylose is low because the ao group forms Schiff base with reducing saccharides. HILICpak VG-0 is the ao column that the recovery ratio of reducing saccharides is improved. By the improvement of the recovery ratio, the sensitivity of the analysis gets higher. Recovery of Mannose (%) Sample : mg/ml each, µl. Lactulose. Epilactose. Lactose m/z 0 (+) m/z () Rare sugar 0 0 Column : Shodex HILICpak VG-0 E Eluent : H O/CH CN/CH OH=//0 Flow rate : 0.mL/ Column temp. : 0 C Sample : 0.% each, 0µL. L-Ribose. D-Psicose. D-Xylitol. D-Tagatose. D-Allose. L-Glucose 0 LC/MS analysis of pantothenic acid and vita C Sample : 00ng/mL each, 0µL. Vita B (Pantothenic acid) as Calcium pantothenate. Vita C (Ascorbic acid) Columns for Polymer-based Hydrophilic Interaction Chromatography (HILIC) (HILICpak) 0 0 Column : Shodex HILICpak VG-0 E Eluent : H O/CH CN/CH OH=//0 Flow rate :.0mL/ Column temp. : 0 C Column : Shodex HILICpak VT-0 D Eluent : 0mM HCOONH aq./ch CN=0/0 Flow rate : 0.mL/ Detector : ESI-MS (SIM) Column temp. : 0 C LC/MS analysis of glyphosate and glufosinate LC/MS analysis of phosphorylated saccharides AMPA (Aomethylphosphonic acid) Sample : µg/ml each, µl m/z 0 () Sample : µm each, µl. Glucose--phosphate (GP). Fructose--phosphate (FP). Glucose--phosphate (GP). Fructose--phosphate (FP) Glufosinate m/z (+) Glyphosate m/z () MPPA (-Methylphosphinicopropionic acid) m/z (+) Column : Shodex HILICpak VT-0 D Eluent : H O/% HCOOH aq./ch CN=0/0/0 Flow rate : 0.mL/ Detector : ESI-MS (SIM) Column temp. : 0 C Column : Shodex HILICpak VT-0 D Eluent : mm HCOONH aq./ch CN=0/0 Flow rate : 0.mL/ Detector : ESI-MS (SIM Negative : m/z 9) Column temp. : 0 C US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

24 Columns for Polymer-based Hydrophilic Interaction Chromatography (HILIC) (Asahipak) Features NHP NHP-0 Suitable for saccharides analysis by hydrophilic interaction chromatography (HILIC) Polymer-based packing material provides excellent chemical stability and imum deterioration over extended time period Easily regenerated by washing in a alkaline solution Also suitable for evaporative light scattering detector, corona charged aerosol detector, and LC/MS Corresponds to USP L Provides higher theoretical plate number compared to NHP-0 series Corresponds to USP L Standard columns Functional Group Shipping Solvent F000 Asahipak NHP-0 B,00 Ao CH CN F000 Asahipak NHP-0 D,00 Ao CH CN F000 Asahipak NHP-0 E,00 Ao CH CN F00 Asahipak NHP-0G A Ao. 0 CH CN F000 Asahipak NHP-0 D,00 Ao CH CN F000 Asahipak NHP-0G A Ao.0 0 CH CN F000 Asahipak NHP-0 E,00 Ao CH CN F000 Asahipak NHP-0G A Ao.0 0 CH CN F000 Asahipak NHP-0 B,000 Ao CH CN F0009 Asahipak NHP-0 D,00 Ao CH CN F000 Asahipak NHP-0 E,000 Ao CH CN F000 Asahipak NHP-LF (line filter) Ao.0 CH CN mm I.D columns [Customized columns] Base Material : Polyvinyl alcohol Functional Group F00 Asahipak NHP-0 B Ao F00 Asahipak NHP-0 D Ao Preparative columns * Preparative columns are made to order. Standard Column F000 Asahipak NHP-0 0E 0, NHP-0 F00 Asahipak NHP-90 0F 0, NHP-0 F00 Asahipak NHP-0G B. 0 US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00

25 Fructooligosaccharide syrup Cyclodextrins Stevioside and rebaudioside A 0 Sample : Fructooligosaccharide syrup,.%, 0µL. Fructose. Glucose. Sucrose. -Kestose. Nystose. -Fructofuranosyl-D-nystose 0 0 Column : Shodex Asahipak NHP-0 E Eluent : H O/CH CN=0/0 Flow rate :.0mL/ Column temp. : C Imidazole dipeptides Sample : 0µL. β-alanine 00µg/mL. -Methyl-L-histidine µg/ml. L-Anserine µg/ml. Histidine µg/ml. L-Carnosine µg/ml. Nitrate (derived from L-Anserine Nitrate) Sample : 0µg/mL each, 0µL. α-cyclodextrin. γ-cyclodextrin. β-cyclodextrin Column : Shodex Asahipak NHP-0 E Eluent : H O/CH CN=0/0 Flow rate :.0mL/ Column temp. : 0 C Simultaneous analysis of water-soluble vitas Sample : 0µL. Vita B 0µg/mL. Nicotinamide 0µg/mL. Vita B 0µg/mL. Nicotinic acid 0µg/mL. Folic acid 0µg/mL. Vita C 0µg/mL 0 0 Ascorbic acid and erythorbic acid O=C HOC O HOC HC HCOH CHOH Sample : 0.0% each, 0µL. Stevioside. Rebaudioside A Column : Shodex Asahipak NHP-0 E Eluent : H O/CH CN=/ Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C Sample : µg/ml each, 0µL. Erythorbic acid. L-Ascorbic acid O=C HOC O HOC HC HOCH CHOH Columns for Polymer-based Hydrophilic Interaction Chromatography (HILIC) (Asahipak) Column : Shodex Asahipak NHP-0 E Eluent : 0mM NaH PO aq./ch CN=0/0 Flow rate :.0mL/ Detector : UV (0nm) Column temp. : 0 C 0 Column : Shodex Asahipak NHP-0 E Eluent : 0mM H PO aq./ch CN=/ Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C 0 0 Column : Shodex Asahipak NHP-0 E Eluent : 0mM NaH PO + 0mM H PO aq. /CH CN=0/0 Flow rate :.0mL/ Detector : UV (nm) Column temp. : 0 C Comparison of saccharides analysis using corona charged aerosol detector Sample : 0µg/mL each, µl. Fructose. Glucose Silica-based amido column (.mm I.D. x 0mm) Silica-based ao column (.mm I.D. x 0mm) A corona charged aerosol detector measures effluents as particles. Accordingly, the baseline will be significantly influenced by the components eluted from the column. NHP series, the polymer based ao columns eliate the components originated from the column, resulting a stable baseline with lower noise level. NHP-0 E (.mm I.D. x 0mm) 0 0 Column : Shodex Asahipak NHP-0 E Silica based ao column from other manufacturer Silica based amide column from other manufacturer Eluent : H O/CH CN=0/0 Flow rate :.0mL/ Detector : Corona charged aerosol Column temp : 0 C (NHP-0 E, Silica based ao column) 0 C (Silica based amide column) LC/TOF-MS analysis of -ao benzoic acid derivatized sugar chains Sample : µl Human IgG (µg protein) AA-labeled N-Glycan * AA : -Aobenzoic acid 0 0 Migration time () Column : Shodex Asahipak NHP-0 D Eluent : (A); 9% CH CN/0.% Formic acid aq. (B); % CH CN/0.% Formic acid aq. Linear gradient; (B%) 0% (0-.), 0-9% (.-0) Flow rate : 0.mL/ Detector : ESI-TOF MS (Polarity : Negative, Full MS range : 000 ) Column temp. : C Data provided by Michihiro Kinoshita Ph.D., Faculty of Pharmacy, Kinki University US: 09 Miratech Drive, Suite 00, San Diego, California 9 E. info@amuzainc.com P..0. F..0.00