Aqueous GPC Eluents and Calibrations

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Eluent Name: Sodium 4-Styrenesulfonate Eluent Polymer Types: NaStS Eluent Composition: 70 % 0.2M NaNO 3, 0.01M NaH 2 PO 4, at ph 7.0; 30 % methanol Column(s): PL Aquagel-OH 40 and PL Aquagel-OH 30 Flow Rate: 1.0 ml/min Sample : 5 to 10 mg/ml Ingredient Amount 1 Sodium nitrate 23.8 g 2 Sodium dihydrogen phosphate 1.68 g 3 Distilled water 1400 ml 4 HPLC grade methanol 600 ml Use 630 ml as excess 5 5M NaOH to ph 7.0 Approx. 2.2 to 2.4 ml required 1 Weigh out two salts and dissolve in water. 2 Adjust to ph 7.0 with NaOH. 4 Degas an excess of HPLC grade methanol for 30 minutes, then add to aqueous phase. 5 Degas for an additional 5 to 10 minutes. Calibration Polymer Type: NaStS homopolymers Number of Calibrants: 10 M p Range: 1,100 to 208,000 1 1,100 1 Higher conc. to clear solvent peaks 2 2,290 1 Higher conc. to clear solvent peaks 3 4,480; 29,500; 148,500 8 mg/ml 4 6,430; 45,100; 208,000 8 mg/ml 5 15,800; 86,500 8 mg/ml Information Supplied By: Peter Iddon Last Updated: 17 September 2003 Last Printed: 03/11/2004 Page 1 of 6

Eluent Name: TRISMA (new) Polymer Types: MPC Eluent Composition: 0.2M NaNO 3, 0.05M TRISMA buffer, at ph 7.0 Column(s): PL Aquagel-OH 40 and PL Aquagel-OH 30 Flow Rate: 0.5 ml/min Sample : 5 to 10 mg/ml Maximum Run Length: 45 minutes Calibration Polynomial: 3rd Order Ingredient Amount 1 Sodium nitrate 34.0 g 2 Tris(hydroxymethyl)aminomethane 6.06 g 3 Tris(hydroxymethyl)aminomethane.HCl 7.88 g The HCl salt 4 Distilled water 2000 ml 5 HCl to ph 7.0 1 Dissolve the three salts in the water. 2 Adjust to ph 7.0 with HCl. 3 Degas for 1 hour under house vacuum. Calibration Polymer Type: PEO homopolymers Number of Calibrants: 10 M p Range: 400 to 78,450 1 400 10 mg/ml 2 3 4 5 6 620; 4,120; 22,450 1,080; 7,100 1,470; 12,600 55,600 78,450 Heating of the higher molecular weight calibrants is required to properly dissolve them. However, excessive heating can degrade the polymers. Information Supplied By: Peter Iddon Last Updated: 17 September 2003 Last Printed: 03/11/2004 Page 2 of 6

Incomplete Data Entered! Eluent Name: TRISMA (old) Polymer Types: MPC Eluent Composition: 0.2M NaNO 3, 0.05M TRISMA buffer, at ph 8-10 Column(s): Pharmacia Biotech Superose 6 Flow Rate: unknown Sample : unknown mg/ml Maximum Run Length: unknown Calibration Polynomial: unknown Order Ingredient Amount 1 Sodium nitrate 34.0 g 2 Tris(hydroxymethyl)aminomethane 6.06 g 3 Tris(hydroxymethyl)aminomethane.HCl 7.88 g The HCl salt 4 Distilled water 2000 ml 1 Dissolve the three salts in the water. 2 Adjust to ph 8-10. 3 Degas for 1 hour under house vacuum. Calibration Polymer Type: PEO homopolymers Number of Calibrants: unknown M p Range: unknown Injection Calibrant M p Value(s) Heating of the higher molecular weight calibrants is required to properly dissolve them. However, excessive heating can degrade the polymers. Heat the PEO solutions to 40 C for 1 hour only. Information Supplied By: Iris Ma Last Updated: 18 November 2003 Last Printed: 03/11/2004 Page 3 of 6

Eluent Name: ph 9 with 30 % methanol Polymer Types: SEMA, GMA Eluent Composition: 70 % 0.2M NaNO 3, 0.01M NaH 2 PO 4, at ph 9; 30 % methanol Column(s): PL Aquagel-OH 40 and PL Aquagel-OH 30 Flow Rate: 1.0 ml/min Sample : 5 to 10 mg/ml Ingredient Amount 1 Sodium nitrate 23.8 g 2 Sodium dihydrogen phosphate 1.68 g 3 Distilled water 1400 ml 4 HPLC grade methanol 600 ml Use 630 ml as excess 5 5M NaOH to ph 9 1 Weigh out two salts and dissolve in water. 2 Adjust to ph 9 with NaOH. 4 Degas an excess of HPLC grade methanol for 30 minutes, then add to aqueous phase. 5 Degas for an additional 5 to 10 minutes. Calibration Polymer Type: PEO homopolymers Number of Calibrants: 10 M p Range: 400 to 78,450 1 400 10 mg/ml 2 3 4 5 6 620; 4,120; 22,450 1,080; 7,100 1,470; 12,600 55,600 78,450 Information Supplied By: Peter Iddon Last Updated: 17 September 2003 Last Printed: 03/11/2004 Page 4 of 6

Eluent Name: Acidic Eluent Polymer Types: Cationic polymers (e.g. DMA, DEA, DPA) Eluent Composition: 0.5 M acetic acid, 0.3 M NaH 2 PO 4, at ph 2 Column(s): ViscoGEL G5000 PWxL and ViscoGEL G2500 PWxL Flow Rate: -1 1 mlmin Sample : Ingredient Amount 1 Acetic Acid 60 g 2 Sodium dihydrogen phosphate 72 g 3 Deionised water 2000 ml 1 Weigh out salts and acid and dissolve in water. 2 Adjust to ph 2 using concentrated HCl if necessary. Calibration Polymer Type: Poly(2-vinylpyridine) homopolymers Number of Calibrants: 8 M p Range: 1,480 to 117,000 1 1,480; 20,900 2 3,200; 50,300 3 6,400; 65,700 4 11,000; 117,000 Information Supplied By: Peter Iddon Last Updated: 3 November 2004 Last Printed: 03/11/2004 Page 5 of 6

Eluent Name: Sugar Methacrylate Monomer Eluent Polymer Types: Polymers based on sugar methacrylate monomers (e.g. LAMA, GAMA) Eluent Composition: 0.1 M NaNO 3, 0.01 M NaH 2 PO 4, at ph 7 Column(s): ViscoGEL alpha 3000 Flow Rate: -1 1 mlmin Sample : Ingredient Amount 1 Sodium nitrate 17.0 g 2 Sodium dihydrogen phosphate 2.4 g 3 Deionised water 2000 ml 1 Weigh out salts and dissolve in water. 2 Adjust to ph 7 using 5 M NaOH. Calibration Polymer Type: Poly(ethylene oxide) homopolymers Number of Calibrants: 10 M p Range: 400 to 78,450 1 400 10 mg/ml 2 620; 4,120; 22,450 3 1,080; 7,100 4 1,470; 12,600 5 55,600 6 78,450 Information Supplied By: Peter Iddon Last Updated: 6 October 2004 Last Printed: 03/11/2004 Page 6 of 6