Simultaneous quantitative analysis of 10 indole compounds, including indole-3-acetic acid (IAA, one of the most important naturally occurring auxins) and some of its metabolites, by high performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) after solid-phase extraction (SPE) was reported for the first time. The analysis was carried out using a reverse phase HPLC gradient elution, with an aqueous mobile phase (containing 0.1% formic acid) modified by methanol. Furthermore, a novel SPE procedure was developed for the pre-concentration and purification of indole compounds using C-18 SPE cartridges. The combination of SPE, HPLC, and LC-MS was applied to screen for the indole compounds present in sugar cane (Saccharum officinarum L.) juice, a refreshing beverage with various health benefits. Finally, four indole compounds were successfully detected and quantified in sugar cane juice by HPLC, which were further unequivocally confirmed by LC-MS/MS experiments operating in the multiple reaction monitoring (MRM) mode.