The biodegradation of crude oil is a consequence of the presence of a specific enzymatic system in the microorganisms selected: the alkane hydroxylase (AlkH). The enzymatic biodegradation has been described since 1994, when the enzyme was first isolated from P. putida (formerly P. oleovorans), but the kinetics of microbial degradation has been weakly considered. We studied and described in this work the kinetics of Arab Light biodegradation, a light crude oil used for gasoline production (46.4% C7–C12 n-alkanes), using two oleophilic strains (Bacillus licheniformis and Pseudomonas putida). Alkanes were extracted from aqueous solutions in the bioreactors by dichloromethane, with a high ratio aqueous:organic volumes (1:0.2 mL) for the amplification of the GC n-alkane signals, and GC spectra were monitored in time over 40 days. Petroleum emulsions were visualized using optical microscopy as a result of biosurfactant segregation, which is necessary for the enzymatic biodegradation of oil by microorganisms. Kinetic analysis in biodegradation of Arab Light (total petroleum hydrocarbons, TPH) exhibits first-order kinetics with 0.098 d−1 and 0.082 d−1 as kinetic coefficients for 8.6 g/L initial crude oil concentration (30 °C), which results in degradation rates of 843 mg/Ld and 705 mg/Ld in B. licheniformis and P. putida, respectively. These results can be applied for oil spill bioremediation, using these microorganisms with the objective of removing contamination by petroleum alkanes.