Lutein is a yellow-coloured carotenoid pigment produced naturally by some plants and microoragnisms. Due to its antioxidant activity, there have been longstanding claims about its health benefits (Granado-Lorencio et al., 2009; Fernández-Sevilla et al., 2010). Their recent demonstrations for eye vision (Christaras et al., 2019; Demmig-Adams et al., 2020), brain health, and cognitive functions (Stringham et al., 2019; Gazzolo et al., 2021), especially in the context of aging, are driving the currently increasing interest for this specific molecule (from 249.7 millions (USD) in 2016 to a projected 491 million by 2029 (MMR, 2023)). Indeed, one of the major difference between lutein (and zeaxanthin) and the other carotenoids is its ability to cross the blood-brain barrier. It can therefore access and accumulate in otherwise unreachable tissues such as the retina and the brain (Stringham et al., 2019). In addition to its antioxidant capabilities, which induces health benefits by fighting off reactive oxygen species, lutein has a light-filtering mechanism for violet-blue color, which contributes to the protection and visual performance of the eye (Stringham et al., 2019). In this regard too, compared to other carotenoids, lutein shows greater filtering effects for short wavelengths, probably due to the polarity of the rings in context with the orientation within the lipid membranes (Junghans et al., 2001; Gazzolo et al., 2021).

Humans, and animals, do not synthesize lutein and need to acquire it through their diets. In the case of a human diet, lutein can found in dark green leafy foods, such as broccoli, lettuce, cilantro, spinach and kale, as well as in yellow-orange fruits and roots, like guava, cashews, sweet potato, corn, peppers, pumpkin and eggs (Ochoa Becerra et al., 2020). However, currently the average dietary intake of lutein in Europeans and North Americans stands at a mere 1.7 mg day−1, while studies show that between 6 and 14 mg day−1 would be needed to reduce the risk of age-related diseases (Hajizadeh-Sharafabad et al., 2019). There is therefore a need to increase the daily lutein dose either by diet modification, or, more surely, by diet supplementation.

While not as renowned as fish oil or magnesium supplementation, lutein-rich diet supplementation are currently available to the general public. To date, all the commercial lutein is extracted from marigold flowers, mainly Mexican/African marigold (Tagetes erecta L.) and French marigold (Tagetes patula L.) cultured in China, India and Mexico (Lin et al., 2015; Ochoa Becerra et al., 2020). The Tagetes genus is a group of plants native to America, from southern United States to South America and different cultivars have been developed for various uses. Its high content of red-yellow pigments, among which lutein stands out (3% of dry petals weight), has led to its cultivation mainly for the production of this value-added compound (Lin et al., 2015; Ochoa Becerra et al., 2020). From a technical point of view, marigolds are cultured seasonally and flowers are harvested from July to October. The harvest is followed by drying and chemical processing of the petals to obtain a lutein rich oleoresin with a final 10.6 kg hectare−1 year−1 productivity (Bosma et al., 2003). Nevertheless, this process suffers some drawback. First, its cultivation requires large amounts of land and water for irrigation. Second, although efforts have been made to develop machinery for harvesting (Willoughby et al., 2000) and processing (Britton et al., 2001) flowers, there is no evidence that these efforts have materialized in commercial agricultural equipment, which means that the work continues to be done manually, with the consequent risks and labor costs. Third, the environmental resources required for this method are substantial, with estimates of 60 m3 of water, 8.2 kg of fertilizers, 556 L of hexane, 11.1 L of ethanol, 1.1 kg of KOH, and 121 MJ of energy needed for every 1 kg of non-esterified lutein produced (Vechpanich and Shotipruk, 2010; Lin et al., 2015). Finally, by shifting the focus from technical to financial consideration, one could state that being a seasonal production with only one harvest per year, lutein intrinsically bares an economical risk. Therefore, marigold's growing conditions, requirements, and the increasing demand for lutein worldwide are encouraging the search for new sources of production.

Consequently, alternatives emerge, namely, a chemical sourcing and a biotechnological sourcing. Although pigments obtained from chemical synthesis are becoming widely rejected and alternatives are being sought, some efforts have been made to synthesize lutein by chemical processes. However, the process involves numerous steps and the yield is very low (between 1 and 5%) (Mayer and Rüttimann, 1980; Khachik and Chang, 2009). For this reason, strategies to produce biologically synthesized lutein are still the most studied. Among them, synthetic biology tools have recently been proposed for the production of lutein from bacteria and yeast, microorganisms that do not produce this carotenoid naturally (Takemura et al., 2021). Microbial fermentation exhibits fast growth rates, making its combination with genetic engineering a promising substitution pathway for the production of value-added compounds (Bian et al., 2021, Bian et al., 2023). A lutein titer of 11 mg L−1 in its free form was obtained from a genetically engineered Escherichia coli (Takemura et al., 2021). On the other hand, engineered Saccharomyces cerevisiae was developed to enable lutein biosynthesis and reached a maximal cell concentration of 19.92 mg L−1 (Bian et al., 2021). Despite the general scientific consensus that products derived from genetically modified organisms are safe for consumption, concerns about their negative effects and low social acceptability hinder its market.

In this landscape, microalgae represent an additional alternative. Indeed, microalgae are postulated as a rich source of carotenoids, offering more favorable cultivation conditions and higher lutein productivity compared to traditional plant crops. They require less water and land, involve less labor intensity, can be cultivated in non-agricultural land, and boast better yield per unit of area, allowing for year-round cultivation (Fernández-Sevilla et al., 2010; Lin et al., 2015). With a wider focus, microalgae are also an attractive source of biomass, natural colourants, and chemical compounds with applications in the food and feed industry, as additives in cosmetics, medicines and nutritional supplements, and as a source of by-products for the formulation of bio-plastics and bio-fuels (Alam et al., 2020; Levasseur et al., 2020). Therefore, microalgal lutein production would not be restricted to a single output product but could enter a more diverse and robust valorization scheme through the concept of biorefinery (Safi et al., 2014).

The content of carotenoids in microalgae, including lutein, has been studied since the 1960s (Iwata et al., 1961). But it is only in the 1980s and 1990s, together with the intensification of research on microalgae cultivation at an industrial level, that the first works on the optimization of carotenoid biosynthesis in microalgae began to appear (Borowitzka et al., 1984; Vonshak, 1985). Since then, the only carotenoid pigments produced industrially from microalgae are astaxanthin and β-carotene. This has been possible due to the capacity of certain microalgae strains to store secondary carotenoids as a survival mechanism. Haematococcus pluvialis and Dunaliella salina, can accumulated up to 4% and 10% (Dry Weight, DW) of astaxanthin and β-caroten, respectively (Pick et al., 2019). These species can accumulate such a large amount of pigments due to cellular mechanisms that respond to stress conditions (Esteban et al., 2015).

However, lutein content among studied microalgal species varies considerably between 0.19 and 0.72% DW (Ho et al., 2014) and only some strains stand out as lutein producers under certain conditions, such as C. vulgaris CS-41 (0.94% DW) (McClure et al., 2019), D. salina (0.88% DW) (Fu et al., 2014) or Parachlorella sp. JD-076 (1.18% DW) (Heo et al., 2018).

As a primary carotenoid, lutein synthesis is linked to biomass growth and there are no known metabolic pathways in microalgae that can lead to lutein sequestration and accumulation in lipid bodies in the chloroplast or cytoplasm, as there are for the accumulation of astaxanthin and β-carotene in some microalgal species (Xie et al., 2021).

In recent years, numerous studies have focused on studying lutein content in microalgae, mainly from the genus Chlorella (C. vulgaris (McClure et al., 2019), C. pyrenoidosa (Sampathkumar and Gothandam, 2019), C. protothecoides (Wei et al., 2008; Ribeiro et al., 2017; Xiao et al., 2018; Shi et al., 2000), C. sorokiniana (Cordero et al., 2011; Chen et al., 2017a), C. zofingiensis (Liu et al., 2014)) and Scenedesmus (S. obliquus (Wiltshire et al., 2000; Ho et al., 2014), S. almeriensis (Sánchez et al., 2008), S. incrassatulus (Flórez-Miranda et al., 2017)), but also on Chlamydomonas reinhartii (Ma et al., 2020b), Muriellopsis sp. (Del Campo et al., 2000) Coccomyxa onubensis (Vaquero Calañas, 2013; Bermejo et al., 2018; Soru et al., 2019), Dunalliela salina (Fu et al., 2014). The majority of these studies focus on understanding how such species respond to changes in culture parameters and how lutein content is affected. The parameters most studied to understand how microalgae adjust the amount of lutein to environmental changes are light (intensity, quality and light-dark cycles), nutrients (mainly nitrogen and carbon), temperature and salinity. Unlike what happens with the accumulation of astaxanthin and β-carotene, the induction of stress by the lack or excess of any of these parameters does not substantially increase the amount of lutein. In fact, in many cases, it reduces it. These stress factors also reduce the capacity to generate microalgal biomass, ultimately affecting overall lutein productivity.

In an economic feasibility comparison between marigold and microalgae lutein production, Lin et al. (2015) suggest that potential microalgal strains must have a lutein content of at least 1% DW to be economically feasible. Furthermore, Xie et al. (2021) calculated the maximal theoretical content a microalgae cell can accumulate, reporting a similar value of 1% DW and presents several limiting factors that must be addressed to achieve higher contents and promote commercial production. Genetic improvement of microalgae has been suggested to increase lutein synthesis and accumulation. The primary improvement mechanism used in microalgae is random mutagenesis. This process selects individuals with desirable characteristics after being subjected to chemical or physical treatments that alter parts of their DNA. While, in certain cases, there is documentation of increased lutein levels, the enhancements achieved fall short of the targeted 1% threshold (Cordero et al., 2011; Chen et al., 2022). Additional methods characterized by targeted modifications, such as knockout of repressor genes or heterologous expression of genes that control the synthesis and cyclization of carotenoid precursors, such as phytoene and lycopene, have yielded interesting results in terms of increase, but still below the 1% (Patel et al., 2022). For example, Rathod et al. (2020) reported a lutein percentage increase of 83% on Chlamydomondas reinhardtii, after an heterologous expression of the phytoene-β-carotene synthase gene from red yeast Xanthophyllomyces dendrorhous. However, the total lutein content was 8.9 mg g−1, still below the target. Additionally, it has then been suggested that future studies should focus on precise targeted DNA modifications using editing techniques, such as CRISPR-Cas9 (Hu et al., 2020). These modifications could focus on increasing the enzymatic activity for esterification of lutein, increasing its resistance to light and ROS damage and providing a first step towards the potential formation of lutein-sequestering lipid bodies (Xie et al., 2021). Although, in recent years, some critical genes for these processes have been identified in plants and microalgae, further advances in microalgae genomics and proteomics are needed to achieve substantial progress.

In the pursuit of harnessing lutein from alternative sources, previous reviews have predominantly focused on assessing lutein content within microalgae, and only a few have highlighted the ability of heterotrophic and mixotrophic growth to increase productivity. Most of them highlight relevant findings and suggest strategies such as inducing oxidative stress, genetic engineering and nutrient concentration variations to increase the cells' lutein content.

For example, Hu et al. (2018) clearly explains the metabolism associated with pigment synthesis when microalgae are grown in heterotrophy and enumerate methods by which enhancements in lutein content have been attained, yet they do not delve into how these approaches affect biomass production and, consequently, lutein productivity. Similarly, reviews from Saha et al. (2020) and Patel et al. (2022) explain in detail the metabolic pathways associated with lutein synthesis, both in the presence and absence of light, as well as the genetic engineering tools reported to increase lutein content, but pay little attention to productivity values. While these efforts have undeniably contributed valuable insights into the biochemical pathways and environmental factors influencing lutein accumulation, the quest for elevating lutein content alone may have reached a plateau.

Despite elaborated adjustments on the culture protocols, the increments in lutein content often remain negligible (Fig. 1), raising a pivotal question: should we continue focusing primarily on content, or is it time to shift our collective attention towards enhancing lutein productivity? The subtle distinction between content and productivity holds immense significance.

Recently, reviews from Zheng et al. (2022a), Fu et al. (2023), and Leong and Chang (2023) have highlighted the importance of heterotrophic and mixotrophic growth modes along with the potential of two-stage cultures to enhance lutein productivity. Furthermore, these reviews offer detailed explanations of three highly relevant topics: different bioreactor systems for microalgae cultivation and lutein production, in-situ accumulation of lutein in fermenters using metabolic engineering, and lutein production at pilot scale. However, these reviews primarily document the high productivity values from the references without deeply analyzing the interplay between lutein content, biomass production, and lutein productivity, which is the focus of the current review.

Hence, in the field of microbial compound production, the distinction between productivity and content is not just a matter of semantics, and lutein production is no exception. While the amount of lutein per unit biomass quantifies the amount of this pigment contained in the cells, productivity encompasses a broader concept. It comprises not only the amount of lutein in the cells but also the capacity of the system to produce these cells efficiently. And efficiency in a process always has one critical aspect: time. Productivity integrates time into the equation, and this aspect is crucial in the context of industrial production, as it directly influences the economic viability of a project for commercial purposes.

As discussed below, when considering the three factors for determining productivity (lutein content, density of biomass obtained and the time required to produce it), we understand that it is not always the species with the highest lutein content that yields the highest productivity. The same is true if we take any one (or even two) of these parameters individually. We could have a microalgae species with the capacity to produce a large amount of lutein-rich biomass. Still, if it takes two months to produce it, its productivity will be lower than that of species with lower capacity but more efficient in time.

In exploring lutein from microalgae, this review endeavors to redirect the spotlight towards lutein productivity.

Our rationale stems from a critical observation: elevating lutein productivity might require a preliminary emphasis on biomass production. By magnifying the biomass output of lutein-producing organisms, we can inherently enhance lutein yield, making the process economically viable.

This perspective shift is both timely and practical, especially in light of the increasing global demand for lutein. As we navigate the landscape of alternative lutein sources, understanding and optimizing lutein productivity could hold the key to unlocking the full potential of microalgae as a sustainable and economically feasible source of this essential nutrient.