# Caroenoid Production from Microalgae: Biosynthesis, Salinity Responses and Novel Biotechnologies
## 5 Future Prospects
### Co-Production of Carotenoids with Value-Added Products from Microalgae  
Lipids and several carotenoids (such as astaxanthin) are secondary metabolites of microalgae under adversity, which means certain stimulating conditions may promote the accumulation of two products simultaneously. In _H. pluvialis_, astaxanthin esterified by FAs is stored in lipid bodies, where TAGs are also positioned. Additionally, the biosynthesis of FAs and TAG has been demonstrated to be correlated with astaxanthin biosynthesis in _H. pluvialis_[121]. By adding 6 uM Cu\({}^{2+}\), the astaxanthin and lipid content was increased by 66.87 and 34.99%, respectively, with carotenogenic and lipogenic genes being upregulated at transcriptional level [122].  
Microalgae also contain biofunctional unsaturated fatty acids. Omega-3 polyunsaturated fatty acids (Omega-3 PUFA) from microalgae, such as eicosapentaenoic acid (EPA. C20:5) and docosahexaenoicenoic acid (DHA, C22:6), have been proved to have various health effects with dietary intake, lowering the risks of neurodegenerative diseases and cardiovascular disorders [123]. _Nannochloropsis oceanica_ oil is rich in EPA, while its zeaxanthin content (30.2 mg/g DW) is relatively high among microalgae species [41], making it a potential species for co-production of zeaxanthin and EPA. _O. aurita_ also contains high concentrations of EPA (28% of total fatty acids), which has been confirmed for large-scale culture in open ponds [47], making it competent for producing both fucoxanthin (18.47 mg/g DW) and EPA.  
In addition, co-production of fucoxanthin and DHA is feasible in microalgae. The marine algae genus _Isochrysis_ is rich in DHA, and it allows easy extraction of DHA and fucoxanthin as it has no cell wall. [PERSON] et al. screened 16 strains of _Isochrysis_ for DHA and fucoxanthin production, and _Isochrysis_ COMP1324 in semi-continuous cultivation excels with productivities as 9.05 and 7.96 mg/L/day of DHA and fucoxanthin, respectively [124]. _I. zhanjiangensis_ accumulated both high content of fucoxanthin (23.29 mg/g DW) and stearidonic acid (SDA, 17.16% of total fatty acid), which is the common precursor of EPA and DHA [46].  
Additionally, both cultivation strategies and efficient extraction methods are worth noting. As zeaxanthin and lipid are soluble in organic solvents, choosing an efficient extraction system, such as hexane: methanol (3:2), can realize the best extraction efficacy for both EPA (36.4 mg/g DW) and zeaxanthin (28.2 mg/g DW), which also avoid the usage of toxic chloroform [41]. As most fucoxanthin was mostly in the hydroalcoholic phase (over 99%), lipid and fucoxanthin can be separated by this biphasic system after the same extraction process with pure ethanol solvent [50].  
The residual biomass after carotenoid extraction of microalgae can be further processed to harvest other by-products. Under nutrient starvation, _D. salina_ accumulated \(\beta\)-carotene and triglycerides up to 30-60% DW [125]. After carotenoid extraction, the residuals can be employed for co-production of biodiesel after transesterification. In addition, the carbohydrate in _D. salina_ can also be employed for bioethanol production [126].