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High-Performance Liquid Chromatographic Analysis of the 2-Chloroprocaine Metabolite, Diethylaminoethanol, in Blood and Serum
  1. Qing C. Meng, Ph.D.,
  2. Thomas H. Kramer,
  3. Pharm. D.,
  4. G. Richard Arthur, Ph.D.,
  5. Philip S. Kim, M.D. and
  6. F. Michael Ferrante, M.D.
  1. From the Department of Anesthesia, The Center for Anesthesia Research, and the Pain Medicine Center, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania.
  1. Reprint requests: F. Michael Ferrante, M.D., Pain Medicine Center, Suite 140 Medical Office Building, Presbyterian Medical Center, 39th & Market Streets, Philadelphia, PA. 19104-2699.


Background and Objectives. 2-Chloroprocaine is rapidly metabolized in the blood to yield 2-chloro-para-aminobenzoic acid (an inactive metabolite) and diethylaminoethanol (DEAE). DEAE possesses local anesthetic activity. The only reported assay for DEAE is a colorimetric method.

Methods. Clinical samples of whole blood and serum were obtained from patients receiving stepped intravenous infusions of 3% 2-chloroprocaine. A high pH-dependent liquid-liquid extraction step with diethyl ether was used to eliminate interfering peaks in high-pressure liquid chromatography (HPLC) analysis. Separation and quantitation were performed using HPLC on a polymeric-reversed phase column with a mobile phase consisting of 10% or 20% acetonitrile (for whole blood or serum analysis, respectively) in 50 mm aqueous sodium phosphate buffer, pH = 11.50. The elution order of DEAE and its analogues was tested to interpret the HPLC separation mechanism.

Results. Extraction recovery of DEAE from whole blood was 67 ± 13.5%, from serum, 71 ± 12.2%, and from water, 75 ± 2.9%. The high pH value of the mobile phase resulted in sharp, well-resolved peaks with run times of approximately 8 minutes using 20% acetonitrile. The lower limit of detection was 5 ng/mL of DEAE from a 1-mL sample. The elution order of DEAE and its analogues indicated that separation was based on the hydrophobicity of the analytes rather than polar group interactions occurring with silica-based stationary phase.

Conclusions. A new, simple and rapid HPLC method for extraction and measurement of DEAE in whole blood or serum samples is reported here.

  • High performance liquid chromatography
  • 2-chloroprocaine
  • metabolite
  • diethylaminoethanol
  • ester-linked local anesthetics.

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