Abstract Aloe fleurentiniorum is a desert plant that is occasionally used to treat wounds by local people of Aridah and Fayfa mountains in Saudi Arabia. However, medicinal value of the plant has not been scientifically established. The purpose of this study was to determine the bioactive phytocomponents contained in the exudate gel (EG) from the leaves of Aloe fleurentiniorum using GC–MS and FT-IR studies, as well as antibacterial assays. Medicinally important bioactive compounds were identified using GC–MS analysis. The bioactive compounds are “pregn-5-ene-3α,20-diol”, “16α-methyl-pregnane-11,20-dione”, “3-hydroxy-(3α,5α)-, ursodeoxycholic acid”, “1-heptatriacotanol”, “allopregnane-7α,11α-diol-3,20-dione”, “D-arabino-hexopyranoside”, “2-octadecenoic acid methyl ester”, “D-ribo-hexose,2,6-dideoxy-3-O-methyl glucosamine”, “N-acetyl-N-benzoyl-alpha-D-glucopyranoside”, “nonyl 1-thio pregnenolone”, “5-cholestene-3-ol, 24-methyl-cholestanol”, and “D-allose hexadecanoic acid methyl ester”. Furthermore, specific groups and their respective chemical compounds were identified via FT-IR spectroscopy studies. The FT-IR spectroscopy of EG showed various functional groups at 3354, 2945, 2832, 2523, 2046, 1707, 1451, 1108, 1031, 880, 737 and 610 cm−1. The FT-IR peaks revealed the likely presence of various compounds such as glycosides, flavonoids, steroids, saponins, flavonoids, amino sugars, cutin and isothiocyanate. Moreover, EG produced a wide range of antibacterial effects on some screened human pathogenic bacteria.