The scope of this work was to contribute to the assessment of PHA production process from mixed microbial cultures and complex waste streams by process simulation and prospective life cycle assessment to support the scale-up of the process. Therefore, three scenarios were constructed based on the valorisation of waste streams available in Bogotá, Colombia. The scenario development methodology involves the definition of goal and scope as stated in the life cycle assessment methodology: ISO 14040: 2006 and 14044: 2006. Experimental data from literature was used to define process parameters and yields. A three-stage configuration was identified as the most common ensemble for the unit operations and unit procedures required to transform waste into PHA. Operational conditions and operational parameters were defined to construct process flow diagrams for each scenario. The scenarios pursued the evaluation of the valorisation of sewage sludge (Scenario 1, S1), organic fraction of solid waste (Scenario 2, S2), and the mixture of the residues (Scenario 3, S3). Each scenario was evaluated from a technical and environmental perspective, and the functional unit for the evaluation was 1,7 kg CODPHA (1 kg of PHA). The process simulation was performed in SuperPRO Designer, while the prospective life cycle assessment was completed in SimaPro. The average overall yields obtained during the simulation were: S1 30.80, S2 7.80, S3 12.52 kg VS per kg of PHA; the scenario with the highest yield was Scenario 2. The result was a prospective environmental assessment that made possible the identification and reduction of gaps in terms of carbon source production, biomass production, PHA production, and cross-cutting aspects for technology transfer.