The Use of Acetone As a Substitute for Acetonitrile in Analysis of Peptides by Liquid Chromatography/electrospray Ionization Mass Spectrometry

The recent worldwide shortage of acetonitrile has prompted interest in alternative solvents for liquid chromatography/mass spectrometry (LC/MS). In this work, acetone was substituted for acetonitrile in the separation of a peptide mixture by reversed-phase high-performance liquid chromatography (RP-HPLC) and in the positive electrospray ionization mass spectrometry (ESI-MS) of individual peptides. On both C12 and C18 stationary phases, the substitution of acetone for acetonitrile as the organic component of the mobile phase did not alter the gradient elution order of a five-peptide retention standard, but did increase peak width, shorten retention times, and increase peak tailing. Positive ESI mass spectra were obtained for angiotensin I, bradykinin, [Leu(5)]-enkephalin, and somatostatin 14 dissolved in both acetonitrile/water/formic acid (25%/75%/0.1%) and acetone/water/formic acid (25%/75%/0.1%). Under optimized ESI-MS conditions, the mass spectral response of [Leu(5)]-enkephalin was increased two-fold when the solvent contained acetone. The substitution of acetone for acetonitrile resulted in only slight changes in the responses of the remaining peptides. A higher capillary voltage was required for optimum response when acetone was used. Compared with acetonitrile/water/formic acid (50/50/0.1%), more interfering species below m/z = 140 were found in the ESI-MS spectra of acetone/water/formic acid (50/50/0.1%).