Ketocarotenoids are high-value pigments used commercially across multiple industrial sectors as colorants and supplements. Chemical synthesis using petrochemical-derived precursors remains the production method of choice. Aquaculture is an example where ketocarotenoid supplementation of feed is necessary to achieve product viability. The biosynthesis of ketocarotenoids, such as canthaxanthin, phoenicoxanthin, or astaxanthin in plants is rare. In the present study, complex engineering of the carotenoid pathway has been performed to produce high-value ketocarotenoids in tomato fruit (3.0 mg/g dry weight). The strategy adopted involved pathway extension beyond β-carotene through the expression of the β-carotene hydroxylase (CrtZ) and oxyxgenase (CrtW) from Brevundimonas sp. in tomato fruit, followed by β-carotene enhancement through the introgression of a lycopene β-cyclase (β-Cyc) allele from a Solanum galapagense background. Detailed biochemical analysis, carried out using chromatographic, UV/VIS, and MS approaches, identified the predominant carotenoid as fatty acid (C14:0 and C16:0) esters of phoenicoxanthin, present in the S stereoisomer configuration. Under a field-like environment with low resource input, scalability was shown with the potential to deliver 23 kg of ketocarotenoid/hectare. To illustrate the potential of this “generally recognized as safe” material with minimal, low-energy bioprocessing, two independent aquaculture trials were performed. The plant-based feeds developed were more efficient than the synthetic feed to color trout flesh (up to twofold increase in the retention of the main ketocarotenoids in the fish fillets). This achievement has the potential to create a new paradigm in the renewable production of economically competitive feed additives for the aquaculture industry and beyond. Significance Ketocarotenoids are high-value pigments used in the food and feed industry to confer color. Aquaculture is a good example, where the addition of carotenoids to the feed is essential for the coloration of trout or salmon flesh, and thus product viability. In this study, complex engineering has been carried out to produce a renewable source of ketocarotenoids for use as feed additives. Production in tomato fruit has enabled the testing of this “generally recognized as safe” material with low-energy minimal bioprocessing in aquaculture trials to demonstrate production, technical, and economic feasibility of the system. This achievement represents a potential paradigm in the bioproduction of specialty and bulk chemicals without our reliance on fossil fuel-derived chemical processes.

Sophorolipids (SLs) are microbial biosurfactants with an important role in industry and a continuously growing market. This research addresses the use of sustainable resources as feedstock for bioproducts. Winterization oil cake (WOC) and molasses are suitable substrates for SLs via solid-state fermentation (SSF). The model proposed herein was established for annually processing 750 t of WOC and comparing three support materials: wheat straw (WS), rice husk (RH), and coconut fiber (CF). Production capacity ranged 325–414 t of SLs per year. Unit Production Cost was 5.1, 5.7, and 6.9 USD/kg SL for WS, RH, and CF production models, respectively, and was slightly lower with other substrates. Financial parameters were CAPEX 6.7 MM USD and OPEX 1.9 MM USD/y, with a NPV, IRR and payback time of 6.4 MM USD, 31% and 3.2 y, respectively. SLs recovery from the solid matrix was the major contributor to operating costs, while fermentation equipment shaped capital costs. Results show that the physical properties (bulk density, WHC) of substrates and supports define process costs beyond substrate purchase costs and process yields in SSF systems. To our knowledge, this is the first attempt to model SLs production via SSF at full scale for the economic valuation of the SSF process.

Taraxacum officinale (L.) Weber ex F.H. Wigg, commonly known as dandelion, is a cosmopolitan and perennial weed, which has medicinal properties. In vitro propagation methods are widely used on plants that have difficulties in cultivation and, consequently, low extraction yields of active metabolites. Thus, callus culture has been considered to be useful for the accumulation of several metabolites. In this study, we aimed to establish an efficient protocol for callus induction and maintenance of T. officinale, for which explant type, carbon source, light conditions, and nine different combinations of plant growth regulators (PGRs), such as 1-naphthaleneacetic acid (NAA) (from 0.05 to 0.5 mg/L) and 6-benzylaminopurine acid (BAP) (from 0.5 to 3.0 mg/L), were evaluated. The results showed that hypocotyls and roots from sterile seedlings are the best sources for callus induction, with 100% of callogenesis at every condition tested, and more than 95% of viability and friability. Complete darkness and a medium supplemented with sucrose at 2.3% (w/v) and 0.5 mg/L of NAA and 0.5 mg/L of BAP were the best conditions for callus induction, showing callus with low organogenesis and high friability. This study provides a basis for future studies on improving large-scale callus propagation and further establishment of suspension culture systems for commercial purposes.

Pentacyclic triterpenes, including lupeol, α- amyrin, and β-amyrin, present a large range of biological activities including anti-inflammatory, anti-cancer, and gastroprotective properties. The phytochemistry of dandelion (Taraxacum officinale) tissues has been widely described. Plant biotechnology offers an alternative for secondary metabolite production and several active plant ingredients are already synthesized through in vitro cultures. This study aimed to establish a suitable protocol for cell growth and to determine the accumulation of α-amyrin and lupeol in cell suspension cultures of T. officinale under different culture conditions. To this end, inoculum density (0.2% to 8% (w/v)), inoculum age (2- to 10-week-old), and carbon source concentration (1%, 2.3%, 3.2%, and 5.5% (w/v)) were investigated. Hypocotyl explants of T. officinale were used for callus induction. Age, size, and sucrose concentrations were statistically significant in cell growth (fresh and dry weight), cell quality (aggregation, differentiation, viability), and triterpenes yield. The best conditions for establishing a suspension culture were achieved by using a 6-week-old callus at 4% (w/v) and 1% (w/v) of sucrose concentration. Results indicate that 0.04 (±0.02) α-amyrin and 0.03 (±0.01) mg/g lupeol can be obtained in suspension culture under these starting conditions at the 8th week of culture. The results of the present study provide a backdrop for future studies in which an elicitor could be incorporated to increase the large-scale production of α-amyrin and lupeol from T. officinale.