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The slow action of fungi is one of the biggest challenges in using entomopathogenic fungi. A promising alternative to reduce the time of action is to combine conidia with extracellular enzymes. This study aimed to characterize the production of Pr1 subtilisin protease and lipases by <i>Beauveria bassiana</i> and <i>Metarhizium anisopliae</i> in different culture media and to evaluate the efficiency of the enzymatic treatment against <i>Aphis gossypii</i> and <i>Spodoptera frugiperda</i>. The isolates were cultivated in five different liquid cultures, and, after 7 days, the culture was filtered and centrifuged, and the activity of the Pr1 and lipases was measured. The fungi cultured in a Luria–Bertani broth medium had the highest activity of proteases and lipases. The mortality of <i>A. gossypii</i> nymphs treated with conidia 7 days after the treatment was 39% (JEF-410), 76.5% (JEF-492), 74.8% (ERL-836), and 70.9% (JEF-214). The <i>B. bassiana</i> JEF-410 supernatant combined with conidia increased the fungal virulence at day 5 and day 6 after treatment. When <i>S. frugiperda</i> larvae were treated with <i>B. bassiana</i> JEF-492 conidia combined with its supernatant, the time of infection was shorter compared to the larvae treated with conidia only. Once the supernatant was incubated at 37 °C, the relative activity decreased from 100% to 80% after 2 h and to 45% after 24 h. The results suggest that the supernatant of entomopathogenic fungi may be formulated and used as a biopesticide in an efficient strategy for the biological control of pests.