Arachodonic acid

The HPLC equipment is a Perkin-Elmer Nelson Model 1022 Plus Chromatograph, connected to a Perkin-Ehner series 200 LC Pump, a LC295 UV/VIS detector, a 7125 BIO injector with a 20-pL loop, and a Canberra Packard Elow Scintillation Analyzer 500 TR Series, with a 0.5-mL flow cell. The system is interfaced with a Compaq Prolinea 5100 computer, using a Canberra Packard ELO-ONE software for system control and data processing. The RP-HPLC columns are Pecosphere Ci8, 5 jam, 30 X 3 mm i.d. (SGE Inc.), with a 5- im C18 guard column (SGE Inc.). Mobile phase is a mixture of methanol water acetic acid (85 15 0.1), and the elution of metabolites is done at a constant flow rate of 0.8 mL/min. Absorbance values are measured at 204 nm. Liquid scintillation cocktail (Ultima Ho M, Canberra Packard) is mixed with the eluent at a 1 2 (eluent(LSC) ratio. Retention times of AEA and arachodonic acid (AA) are 3.2 and 4.5 min, respectively. The concentration of AA is calculated from radioactivity of peak area, and FAAH specific activity is expressed as pmol AA released/min/mg protein. 

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