Determination of endogenous concentrations of N1-methylnicotinamide in human plasma and urine by high-performance liquid chromatography

Anal Biochem. 1990 May 15;187(1):160-5. doi: 10.1016/0003-2697(90)90435-c.


A high-performance liquid chromatographic assay for the determination of endogenous plasma and urine concentrations of N1-methylnicotinamide was developed. N1-Methylnicotinamide and N1-ethylnicotinamide (internal standard) are reacted with acetophenone in a strong base at 0 degree C, formic acid is added, and the reaction mixture is heated in a boiling water bath, resulting in the formation of fluorescent derivatives. These derivatives were chromatographed on a C18 reverse-phase column using a mobile phase of acetonitrile-triethylamine and 0.01 M heptanesulfonic acid adjusted to pH 3.2. Fluorescent detection was achieved using 366-nm excitation and 418-nm emission filters. Precision and accuracy were generally greater than 90%, interfering peaks did not cochromatograph, and the limit of quantification was 2 ng/ml in plasma using a 0.2-ml sample. The method was used to examine the concentrations of endogenous N1-methylnicotinamide in the plasma of 36 subjects with various pathology. The mean concentration was 18 ng/ml and the range was 6.2 to 116.7 ng/ml. The assay represents a marked improvement on previous methods and is suitable for routine clinical monitoring.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetophenones / metabolism
  • Adolescent
  • Adult
  • Aged
  • Aging / metabolism
  • Chromatography, High Pressure Liquid / methods*
  • Female
  • Fluorescent Dyes
  • Humans
  • Kidney Diseases / blood
  • Liver Cirrhosis / blood
  • Male
  • Middle Aged
  • Niacinamide / analogs & derivatives*
  • Niacinamide / blood
  • Niacinamide / urine
  • Reproducibility of Results


  • Acetophenones
  • Fluorescent Dyes
  • Niacinamide
  • acetophenone
  • N-methylnicotinamide