Page:NIOSH Manual of Analytical Methods - 5002.pdf/2

 REAGENTS:


 * 1) Warfarin, 99%.*
 * 2) Methanol, HPLC grade.*
 * 3) Water, HPLC grade.
 * 4) Phosphoric acid, 85%.*
 * 5) Calibration stock solution, 2 mg/mL.* Dissolve 20.0 mg Warfarin in 10 mL methanol.
 * 6) Mobile phase component, 0.0025 N phosphoric acid. Dissolve 0.06 mL 85% phosphoric acid in 1 L water.

  * See SPECIAL PRECAUTIONS.  EQUIPMENT:


 * 1) Sampler: 37-mm diameter PTFE membrane filter, 1-µm pore size, and cellulose backup pad in two-piece filter holder held together with tape or shrinkable band.
 * 2) Personal sampling pump, 1 to 4 L/min, with flexible connecting tubing.
 * 3) High pressure liquid chromatograph with 20 to 50-µL injection loop; 25 to 30 cm, 10-µm C18 column; UV absorption detector at 280 nm; and integrator.
 * 4) Jars, squat form, 60-mL, ointment, with PTFE film gaskets and screw caps.
 * 5) Tweezers.
 * 6) Pipet, 5-mL, and pipet bulb.
 * 7) Syringes, microliter.
 * 8) Volumetric flasks, 10-mL and 1-L.

SPECIAL PRECAUTIONS: Methanol is flammable and toxic. Use in fume hood and away from ignition sources.

Warfarin is toxic and can be absorbed through the skin.

Phosphoric acid is corrosive. Use gloves, goggles and other appropriate equipment to prevent eye contact and repeated or prolonged skin contact. Wash skin with water and change clothes if contact occurs.

SAMPLING:
 * 1) Calibrate each personal sampling pump with a representative sampler in line.
 * 2) Sample at an accurately know flow rate between 1 and 4 L/min for a sample size of 200 to 1000 L. Do not exceed a total loading of 2 mg on the filter.

SAMPLE PREPARATION: Transfer filter with tweezers to jar. Add 5.0 mL methanol. Cap the jar. Swirl to wet filter thoroughly.  CALIBRATION AND QUALITY CONTROL: Calibrate daily with at least six working standards covering the range 4 to 120 µg Warfarin per sample.  Add aliquots (2 to 60 µL) of calibration stock solution to 5 mL methanol in ointment jars. Analyze the working standards together with the samples and blanks (steps 7 and 8). NOTE: Duplicate injections should differ by no more than 3% in peak area.</li> <li>Prepare a calibration graph (peak area vs. µg Warfarin).</li> </ol> <li>Determine recovery at least once in the calibration range for each lot of filters. Prepare three filters at each of five levels plus three media blanks.</li> <ol style="list-style-type:lower-alpha;"> <li>Inject known volumes of calibration stock solution onto blank filters with a microliter syringe. Include media blanks.</li> </ol> </ol>