Sustainable Production of Bio-Based Geraniol: Heterologous Expression of Early Terpenoid Pathway Enzymes in <i>Chlamydomonas reinhardtii</i>

Geraniol is a monoterpene alcohol with a rose-like aroma, used in food and cosmetics and for its anti-inflammatory, antibacterial, and insect-repellent properties. Geraniol is commonly chemically synthesized from petroleum-based sources in a highly energy-demanding process with a large carbon footprint. Alternatively, geraniol can be derived from plant-based essential oils but with relatively low yields and limitations from seasonal cultivation. Here, a sustainable geraniol biosynthesis alternative was established in the photosynthetic green microalga <i>Chlamydomonas reinhardtii</i>. Three enzymes─geraniol synthase from <i>Catharanthus roseus</i> (<i>Cr</i>GES), geranyl diphosphate synthase from <i>Lithospermum erythrorhizon</i> (<i>Le</i>GPPS), and a modified 1-deoxy-d-xylulose-5-phosphate synthase from <i>Salvia pomifera</i> (<i>Sp</i>DXS)─were strategically redesigned for high expression from the algal nuclear genome. Various enzyme combinations and subcellular localizations were tested, resulting after 48 h in up to 1 mg geraniol/L (corresponding to 1.8 mg/g of dry weight) secreted into the culture medium. This work demonstrates a promising route for sustainable, CO₂-based production of geraniol in microalgae and provides a foundation for further optimization.