Industrial scale-up of microalgal cultures is often a protracted step prone to culture collapse and the occurrence of unwanted contaminants. To solve this problem, a two-stage scale-up process was developed – heterotrophically Chlorella vulgaris cells grown in fermenters (1^st stage) were used to directly inoculate an outdoor industrial autotrophic microalgal production unit (2^nd stage). A preliminary pilot-scale trial revealed that C. vulgaris cells grown heterotrophically adapted readily to outdoor autotrophic growth conditions (1-m³ photobioreactors) without any measurable difference as compared to conventional autotrophic inocula. Biomass concentration of 174.5 g L⁻¹, the highest value ever reported for this microalga, was achieved in a 5-L fermenter during scale-up using the heterotrophic route. Inocula grown in 0.2- and 5-m³ industrial fermenters with mean productivity of 27.54 ± 5.07 and 31.86 ± 2.87 g L⁻¹ d⁻¹, respectively, were later used to seed several outdoor 100-m³ tubular photobioreactors. Overall, all photobioreactor cultures seeded from the heterotrophic route reached standard protein and chlorophyll contents of 52.18 ± 1.30% of DW and 23.98 ± 1.57 mg g⁻¹ DW, respectively. In addition to providing reproducible, high-quality inocula, this two-stage approach led to a 5-fold and 12-fold decrease in scale-up time and occupancy area used for industrial scale-up, respectively.

Optically pure lactic acid (LA) is needed in PLA (poly-lactic acid) production to build a crystalline structure with a higher melting point of the biopolymer than that of the racemic mixture. Lignocellulosic biomass can be used as raw material for LA production, in a non-food biorefinery concept. In the present study, genetically engineered P. acidilactici ZP26 was cultivated in a simultaneous saccharification and fermentation (SSF) process using steam pretreated softwood solids as a carbon source to produce optically pure D-LA. Given the low concentrations of identifiable inhibitory compounds from sugar and lignin degradation, the fermentation rate was expected to follow the rate of enzymatic hydrolysis. However, added pretreated solids (7% on weight (w/w) of water-insoluble solids [WIS]) significantly and immediately affected the process performance, which resulted in a long lag phase (more than 40 h) before the onset of the exponential phase of the fermentation. This unexpected delay was also observed without the addition of enzymes in the SSF and in a model fermentation with glucose and pretreated solids without added enzymes. Experiments showed that it was possible to overcome the extended lag phase in the presence of pretreated softwood solids by allowing the microorganism to initiate its exponential phase in synthetic medium, and subsequently adding the softwood solids and enzymatic blend to proceed to an SSF with D-LA production.

Recycling of textile fibers is a mandatory step in the life-cycle of fabrics to avoid incineration or landfilling. After fiber-to-fiber recycling, cellulose fiber saccharification opens the door to the production of biochemical building blocks through microbial fermentation. This study documents for the first time the production of optically pure D-lactic acid (D-LA), one of the precursors of polylactic acid (PLA), from post-consumer waste viscose. Enzymatic hydrolysis was applied to white and colored waste viscose to produce two glucose-rich solutions that were used as carbon source for the genetically modified Pediococcus acidilactici ZP26. Afterward, batch and fed-batch cultivations were conducted with very similar results for the control and both waste viscose hydrolysates (white and colored). The strain was able to produce D-LA without any signs of inhibition by possibly toxic compounds present in the recycled textiles (additives, dyes, and other contaminants). The highest D-LA concentration and yield was reached in the batch reactors with colored waste viscose at 42.4 g/L and 0.97 g/g, respectively. In the fed-batch cultivations, the best result was achieved with the white waste solution, with 59.5 g of D-LA (40.8 g/L), and a yield on total glucose of 0.70 g/g. Notwithstanding the need for more research in the utilization of saccharified waste viscose as carbon source for bioprocesses, this is an important proof of concept of an application of streams from tertiary recycling of cellulosic fibers in the production of platform biochemicals. Graphical Abstract