High-performance liquid chromatographic analysis of the 2-chloroprocaine metabolite, diethylaminoethanol, in blood and serum

Background and Objectives. 2-Chloroprocaine is rapidly metabolized in the b lood to yield 2-chloro-para-aminobenzoic acid (an inactive metabolite) and diethylaminoethanol (DEAE). DEAE possesses local anesthetic activity. The o nly reported assay for DEAE is a colorimetric method. Methods. Clinical sam ples of whole blood and serum were obtained from patients receiving stepped intravenous infusions of 3% 2-chloroprocaine. A high pH-dependent liquid-l iquid extraction step with diethyl ether was used to eliminate interfering peaks in high-pressure liquid chromatography (HPLC) analysis. Separation an d quantitation were performed using HPLC on a polymeric-reversed phase colu mn with a mobile phase consisting of 10% or 20% acetonitrile (for whole blo od or serum analysis, respectively) in 50 mm aqueous sodium phosphate buffe r, pH = 11.50. The elution order of DEAE and its analogues was tested to in terpret the HPLC separation mechanism. Results. Extraction recovery of DEAE from whole blood was 67 +/- 13.5%, from serum, 71 +/- 12.2%, and from wate r, 75 +/- 2.9%. The high PH value of the mobile phase resulted in sharp, we ll-resolved peaks with run times of approximately 8 minutes using 20% aceto nitrile. The lower Limit of detection was 5 ng/mL of DEAE from a 1-mL sampl e. The elution order of DEAE and its analogues indicated that separation wa s based on the hydrophobicity of the analytes rather than polar group inter actions occurring with silica-based stationary phase. Conclusions. A new, s imple and rapid HPLC method for extraction and measurement of DEAE in whole blood or serum samples is reported here.