Selectide Corporation, a subsidiary of Hoechst Marion Roussel, Inc., 1580 E. Hanley Blvd., Tucson, Arizona 85737.

Tel: (520) 544-5850. Fax: (520) 575 8283. Email: marcel.patek@hmrag.com

^‡Biopharm, Research Institute of Biopharmacy and Veterinary Drugs, 254 49 Jílové near Prague, Czech Republic.

Published in: Tetrahedron Lett., 1998, 39, 753-756.

Shortly:

Ionselective electrode (ISE) technique represents another noninvasive method for monitoring and quantitative determination of basic functionalities on solid support. ISE method is based on complete protonation of basic functionalities after treatment of the resin with a large excess of 1% HClO₄. After thorough washing with water, the bound anion is eluted with a suitable base providing an easily detectable perchlorate anion which is quantified potentiomentrically with perchlorate ionselective electrode. Limitations of this technique include the requisite use of watercompatible solid supports and basic functionalities possessing a pK_HB+ > 7.

Experimental:

Materials and Sample Preparation. All materials were obtained from commercial suppliers and used without further purification. Potassium perchlorate, ammonium sulfate, perchloric acid (70%), and sodium bicarbonate were purchased form Aldrich. TentaGel S NH2 (Rapp Polymere, Tübingen, Germany), MBHA (Advanced ChemTech), and (Aminomethyl)polystyrene (Bachem California) resins were used in described experiments. Deionized water (24 mS) was used for the preparation of standards as well as for all measurements. Perchlorate ion selective electrode (doublejunction with BNC connector) was purchased from ColeParmer and connected to pH/mV Meter (F11) from Beckman. Solid phase syntheses of single compounds as well as monitoring steps involving treatment with 1% HClO₄ followed by multiple washings were performed in polypropylene syringes equipped with polyethylene frits.

Stock solutions:

Solution of perchloric acid in methanol (Solution A)

0.1 M HClO₄ (1%)

· HClO₄ : Mw 100.46

Dilute 1 mL of aqueous 70% HClO₄ in 70 mL of methanol (total volume). Note: Perchloric acid is explosive in contact with reductants and is a strong oxidizing agent.

Solution of base with ionic strength adjustor (ISA) (Solution B)

0.024 M NaHCO₃ + 0.33 M (NH₄)₂ SO₄, I = 1 M

· Mw [NaHCO₃]: 84.01; Mw [(NH₄)₂SO₄]: 132.14

Dissolve 2.02 g (0.024 mol) of NaHCO₃ and 43.61 g (0.33 mol) of (NH₄)₂SO₄ in 1L of deionized water.

Standard Solutions:

Standard solutions of known concentration containing ion strength adjustor are used to construct a calibration curve. By serial dilution of the 0.01 M KClO₄ in Solution B, prepare, e.g., 0.08, 0.16, 0.24, 0.32, 0.40, 0.56, and 0.64 mM perchlorate standards.

Procedure for direct determination of an amine concentration on the TentaGel resin:

Sample Preparation and Measurement:

Assuming an amount of 25 mg of the TG resin (0.26 mmol/g) and final volume of the solution to be 25 mL, following protocol provides reliable results with the relative error 3-5%.

1. Wash resin with MeOH (5 x 1 mL).

2. Wash resin with 1% HClO₄ in MeOH (stock Solution A, 2 x 1 mL).

3. Wash out excess of HClO₄ with deionized water (5 x 1 mL).

4. Wash resin with 0.024 M NaHCO₃ + 0.33 M (NH ₄)₂SO₄ (Solution B, 5 x 1 mL).

and collect all washings into 25 mL volumetric flask. Dilute washing with Solution B to the exact volume of 25 mL.

Measurement of perchlorate concentration:

1. Rinse the electrode with deionized water and blot dry.

2. Place the most dilute solution of standard on the magnetic stirrer and stir at a constant rate. Lower the electrode tip into the solution, wait for reading to stabilize and record the mV reading.

3. Repeat steps 1. and 2. for other concentrations of the standard solutions. For quick concentration estimate, only two standards need to be measured. For more precise measurements, set of 5-7 standards should be measured.

6. Rinse the electrode with deionized water and blot dry.

7. Place the sample (15 mL) in a scintillation vial on the magnetic stirrer and stir at constant rate. Lower the electrode tip into the solution, wait for reading to stabilize and record the mV reading.

8. Use Excel worksheet (ise.xls) to calculate the concentration of measured sample. You can download and try the Excel worksheet - ise.zip (16.7 kB). After the transfer, extract files from ZIP archive and run Excel. NOTE: In order to extract files from archive, you will need pkunzip.exe utility.

Note: Since the electrode potential depends on temperature, perform both the standard and sample measurements within the short period of time to eliminate temperature fluctuations. The temperature of both sample solution and standard solutions must be the same.