BUT OX YAC ETIC IN UR INE: ME TH OD 8316, Issue 1, dated 1 5 Ma rch 2003 - Page 2 of 5 REAGENTS:
EQUIPMENT:
1. Te trabu tylam m onium hydrogen sulfate (C 16H 36NAHSO 4). 2. Potassium dihydrogen phosphate. 3. Potassium hydroxide. 4. Phosp horic acid, 85% .* 5. W ater, deionized. 6. C 16H 36NAHSO 4 in phosphate buffer: 0.2 M KH 2PO 4 and 0.1 M C16H 36NAHSO 4 in deionized water, made pH 6 with 85% H 3PO 4 or 10 M KOH. 7. Me thylene chloride.* 8. PF B brom ide, 2,3 ,4,5 ,6-penta fluorobenzyl brom ide.* 9. 2-Propa nol.* 10. To luene.* 11. Isopropano l-toluene m ixture, 1:1 (v/v). 12. PFB-BAA, pentafluorobenzyl ester of buto xyacetic ac id, MW 312 .24 am u. Synthesized ac cording to Groes ene ken [5], purified by RP-HPLC, and analyzed to one m ajor p eak by GC/M S. Dens ity 1.359 g /m L GC-MS (EI) fragmentation pattern; 240, 181, 131, 87, 73, 51amu. 13. PFB-BAA stock standard solution, 680 :mol/L. Dissolve 21 ± 0.1 mg of PFBBAA in 100 mL of 1:1 isopropanol-toluene using an am ber 1 00-m L volum etric flas k. Store at 0 to 5 / C. 14. BAA, Butoxyacetic acid, >99%. 15 Urine from une xpo sed volunteers , store d at -20 to -15 oC for a maximum of 2 weeks. 16. BAA-in-urine stock solution, 4000 :m ol/L. Dissolve 53 ± 1 mg of BAA in 100 mL of urine. Dilute with urine from unexposed volun teers for prepa ration of the qua lity control samples. 17 Argon with 5% methane. 18 Helium.
1. Gas chromatograph with 63Ni electron capture detector, temperature programm ing, splitless injection port with purge, and autoinjector. The detector makeup gas was 5% m ethane in argon at 60 mL/m in. 2. Polypropylen e bo ttles, 30- and 250 -m L with screw caps. 3. Graduated cylinders, 100- and 250-mL. 4. Shipping con tainer, polystyren e foa m , with dry ice. 5. Culture tubes, 16-mm x 100-mm , with teflonlined screw caps, disposable. 6. Disposable serological pipettes , 0.2 -m L in 0.001-m L increm ents, and 2-m L in 0.1-mL increments. Micropipette, 10-:L. 7. Alu m inum foil. 8. Tub e rotator. 9. Centrifuge, 3000 rpm. 10. Evaporator, nitrogen purge-type, with 30 / C heating bath. 11. Vortex m ixer. 12. Ultrasonic water bath, room tem perature. 13. Autoinjector vials, amber with crimp caps. 14. Ba lance, five -decim al-place analytic al. 15. Volumetric flasks, amber, 10- and 100-mL.
- See SPECIAL PRECAUTIONS
SPECIAL PRECAUTIONS: See material safety data sheets. Urine samples may contain a number of bacte rial and viral agents, including hepatitis B virus, a nd should be handled using Biosafety Level 2 prac tices, containm ent equipm ent and fa cilities [CD C & NIH , Biosafety in Microbiological and Biomedical Laboratories, 3rd ed., H HS P ublication No . (C DC) 93 -8395 (1 993)]. Pe nta fluorobenzyl bromide is volatile compound, a powerful lacrimator, and a suspected carcinogen. Methylene chloride, isopropanol, and toluene are NFPA level 3 fire hazards. Phosphoric acid is highly corrosive.
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
