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As an easily obtained organic waste, by-product acetic acid could be an appropriate co-substrate with blue algae wastes (increase C/N ratio of substrates) for co-fermentation of PHA production. However, there are still acrylic acid and other chemicals in by-product acetic acid, which could cause severe inhibition for fermenting microorganisms during PHA production process. The current study represented that alkali pretreatment (pH level of 12) is a more favorable method compared with thermal pretreatment (80 ℃ for 30 min) for breaking cell walls of blue algae. It seemed that there was no synergistic effect of the combination of thermal and alkali pretreatment methods (temperature of 80 ℃ and pH level of 12). Optimal parameters during electro-fenton process for removal of inhibitors in by-product acetic acid were under current of 0.5 A, pH level of 3 and reaction time of 120 min. Both the highest dry weight of PHA and PHA concentration were achieved by applying blue algae and by-product acetic acid (after pretreatment) as co-substrates (mixed ratio of 3:1, stirring speed of 200 r/min, 24 h), indicating that using by-product acetic acid (after pretreatment) as co-substrate could increase C/N ratio and promote PHA production successfully. The current study could offer new insights for improving PHA production by co-fermentation. [Display omitted] • Alkali pretreatment is a more favorable method compared with thermal pretreatment. • There was no synergistic effect of the combination of thermal and alkali pretreatment. • Optimal parameters for removal of inhibitors in by-product acetic acid were determined. • By-product acetic acid (after pretreatment) as co-substrate could promote PHA production. [ABSTRACT FROM AUTHOR]