| Summary: | <i>Alternaria alternata</i> isolates C1, S1, and X3 were isolated respectively from the weeds <i>Convolvulus arvensis</i>, <i>Sonchus oleraceus</i>, and <i>Xanthium strumarium</i> in Algiers during 2016 and identified by morphological and molecular analyses. The aim of this investigation was to chemically characterize the exometabolome of these fungi and to evaluate the myco-herbicidal potential of their culture filtrates, crude extracts, or fractions towards target weeds. Results revealed a great heterogeneity in the biochemical profiles of the exometabolome with the remarkable presence of two compounds: tenuazonic acid (TeA) and triprenyl phenol-7 (SMTP-7). To the best of our knowledge, SMTP-7—found in all the isolates—as well as 12-methoxycitromycin detected in the culture filtrate of isolate C1, have never been reported to be produced by <i>A. alternata</i>. Some fractions of isolates C1 and S1 showed symptoms (necrosis and chlorosis) on the detached leaves of <i>C. arvensis</i> and <i>S. oleraceus</i>, respectively with up to 100% phytotoxic effect at low concentration. In conclusion, biochemical characterization revealed great difference of C1, S1, and X3 exometabolome that is likely to explain the difference in their phytotoxic activity. Some fractions (d<sub>1</sub>, e<sub>1</sub>, h<sub>1</sub>, i<sub>1</sub>, a<sub>2</sub>, and f<sub>2</sub>) of isolates C1 and S1 of <i>A. alternata</i> caused severe necrosis and chlorosis on the injured detached leaves of <i>C. arvensis</i> and <i>S. oleraceus,</i> respectively.
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