Principles of green engineering* Challenges of applying biological CCU I Inherently non-hazardous and safe. The use H2/O2/syngas presents explosion safety challenges to large-scale production. M Minimize material diversity. Less of a biological problem. P Prevention instead of treatment. Bio-wastes are inevitable in fermentation. R Renewable material and energy inputs. Concentration, composition, temperature and pressure of CO2 source have direct impact on organismal growth and productivity. The same applies to energy source (e.g., light intensity and wavelength etc). O Output-led design. Design of biological system is not trivial and requires sound knowledge at both molecular and system level. Robust genetic tool is lacking for modification of some organisms. V Very simple. Biological system is inherently complex, highly integrated and regulated. E Efficient use of mass, energy, space & time. Energy and carbon source are channelled into cell growth and biomass accumulation, instead of chemical production. Low productivity is an issue. Biological membrane could be a barrier to mass/energy transfer. Some enzymes display promiscuous activities (moonlighting). Maintaining strict anoxia for anaerobic cultivation, sparging, and cell stirring can be costly and energy intensive.  M Meet the need. Less of a biological problem. E Easy to separate by design. Most organisms or enzymes are not tolerant to solvents used in product separation. N Networks for exchange of local mass & energy. Less of a biological problem. T Test the life cycle of the design. Less of a biological problem. S Sustainability throughout product life cycle. Less of a biological problem. *Adapted from [126]

Table 6: Challenges of applying biological CCU, identified through principles of green engineering.