A simplified gas chromatographic method for measuring quaternary ammonium compounds has been developed and used to measure the serum concentration of pyridostigmine in human beings. Pyridostigmine is present in the serum within 1 hour after oral administration and reaches a peak at 2 hours. Results in several patients suggest that the serum concentration achieved is related to the size of the dose and that there is a relationship between serum concentration and clinical response.