A validated HPLC method for the determination of pyridostigmine bromide, acetaminophen, acetylsalicylic acid and caffeine in rat plasma and urine.
A method was developed for the separation and quantification of the anti-nerve agent pyridostigmine bromide (PB; 3-dimethylaminocarbonyloxy-N-methyl pyridinium bromide), the analgesic drugs acetaminophen and acetylsalicylic acid, and the stimulant caffeine (3,7-dihydro-1,3,7-trimethyl-1-H-purine-2,6-dione) in rat plasma and urine. The compounds were extracted using C(18) Sep-Pak(R) cartridges then analyzed by high performance liquid chromatography (HPLC) with reversed phase C18 column, and UV detection at 280 nm. The compounds were separated using gradient of 1-85% acetonitrile in water (pH 3.0) at a flow rate ranging between 1 and 1.5 ml/min in a period of 14 min. The retention times ranged from 8.8 to 11.5 min. The limits of detection were ranged between 100 and 200 ng/ml, while limits of quantitation were 150-200 ng/ml. Average percentage recovery of five spiked plasma samples were 70.9+/-9.5, 73.7+/-9.8, 88.6+/-9.3, 83.9+/-7.8, and from urine 69.1+/-8.5, 74.5+/-8.7, 85.9+/-9.8, 83.2+/-9.3, for pyridostigmine bromide, acetaminophen, acetylsalicylic acid and caffeine, respectively. The relationship between peak areas and concentration was linear over range between 100 and 1000 ng/ml. The resulting chromatograms showed no interfering peaks from endogenous plasma or urine components. This method was applied to analyze these compounds following oral administration in rats.
Abu-Qare
A W
AW
Department of Pharmacology and Cancer Biology, Duke University Medical Center, PO Box 3813, Durham, NC 27710, USA.
Abou-Donia
M B
MB
D000082:Acetaminophen / Q000032:analysis* / Q000097:blood / Q000652:urine; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000032:analysis* / Q000097:blood / Q000652:urine; D001241:Aspirin / Q000032:analysis* / Q000097:blood / Q000652:urine; D002110:Caffeine / Q000032:analysis* / Q000097:blood / Q000652:urine; D002138:Calibration; D000697:Central Nervous System Stimulants / Q000032:analysis* / Q000097:blood / Q000652:urine; D002851:Chromatography, High Pressure Liquid; D011729:Pyridostigmine Bromide / Q000032:analysis* / Q000097:blood / Q000652:urine; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results
D000894:Anti-Inflammatory Agents, Non-Steroidal; D000697:Central Nervous System Stimulants; D000082:Acetaminophen; D002110:Caffeine; D011729:Pyridostigmine Bromide; D001241:Aspirin