TRICHLOROACETIC ACID IN URINE: METHOD 8322, Issue 1, dated 17 April 2015 - Page 2 of 5
REAGENTS:
EQUIPMENT:
1. Sodium trichloroacetate [CAS #650-51-1], 97% purity or greater* 2. Boron trifluoride-methanol solution, 14%* 3. Toluene, ACS reagent grade or better* 4. Sodium sulfate, anhydrous, granular; reagent grade or better 5. Acetic acid, glacial; reagent grade or better* 6. Acetone, reagent grade or better* 7. Methanol, reagent grade or better* 8. Helium, purified 9. Nitrogen, Ultra High Purity or P5 10. Water, ASTM Type II [14]
1. Centrifuge tubes, polypropylene, ~15-mL, with screw caps, or other suitable container for specimen collection and storage 2. Gas chromatograph with electron capture detector, autosampler, data collection system and column (page 8322-1) 3. Microliter syringes, various sizes 4. Volumetric flasks, glass, various sizes 5. Adjustable pipettor with disposable plastic tips, 0.1 to 1-mL 6. Disposable Pasteur transfer pipettes, 15 and 23 cm 7. Culture tubes, 13 mm x 100 mm (~8 mL), with PTFE-lined caps 8. Vortex mixer 9. Glass wool 10. Vials, autosampler, glass, 2-mL with caps 11. Oven, capable of maintaining 60 ºC
SOLUTIONS: 1. 1:3 glacial acetic acid:deionized water 2. 1:1 acetone:methanol
- See SPECIAL PRECAUTIONS
SPECIAL PRECAUTIONS: Standard precautions should always be used when handling bodily fluids and/or extracts of bodily fluids [15]. Handle urine specimens and urine extracts using powder-free latex or nitrile gloves. Acetic acid, acetone, toluene, and methanol are flammable; handle with care and use in a chemical fume hood. Handle all chemicals using the required safety precautions. Reagents with manufacturer expiration dates should be observed. The Environmental Protection Agency (EPA) has designated TCAA as a known mouse carcinogen and a possible human carcinogen [16]. SAMPLING: 1. Collect at least 10 mL of urine in ~15-mL polypropylene tubes or other suitable container. NOTE: Because of the relatively lengthy half-life values of TCAA, ACGIH recommends sampling at the end of shift at the end of workweek [1]. 2. Freeze the urine and ship in dry ice in an insulated container. Reminder: commercial shippers have special labeling requirements for packages containing biological samples and dry ice. SAMPLE PREPARATION: 3. Thaw urine specimens, bring to room temperature, and mix thoroughly. 4. Place 200 µL of urine specimen in an 8-mL glass culture tube with a PTFE-lined cap. 5. Add 0.5 mL 14% boron trifluoride in methanol; cap and mix. 6. Heat in oven at 60 ºC for a minimum of 1.5 hr (maximum 2.5 hr). 7. Cool to room temperature and then add 2.0 mL toluene. 8. Vortex or shake vigorously for 1 min. 9. After the layers separate, transfer the upper toluene layer to a drying column containing anhydrous sodium sulfate. The drying columns are prepared in 15-cm Pasteur pipettes with a glass wool plug and about 200 to 300 mg anhydrous sodium sulfate, sufficient to form a bed depth of ~1 cm. 10. Collect the eluate in a 2-mL GC vial. Cap vial.
NIOSH Manual of Analytical Methods (NMAM), Fifth Edition
