Microalgae have long been used for the commercial production of natural colorants such as carotenoids and chlorophyll. Due to the rising demand for carotenoids and other natural products from microalgae, strategies to increase production efficiency are urgently needed. The production of microalgal biorefineries has been limited to countries with moderate climates. For countries with cooler climates and less daylight, methodologies for the efficient production of microalgal biorefineries need to be investigated. Algal strains that can be safely consumed as whole cells are also attractive alternatives for developing as carotenoid supplements, which can also contain other compounds with health benefits. Using such strains helps to eliminate the need for hazardous solvents for extraction and several other complicated steps. In this study, the mesophilic green alga Chlamydomonas reinhardtii was employed to study the effects of cold stress on cell physiology and the production of pigments and storage compounds. The results showed that temperatures between 10 and 20 °C induced carotenoid and chlorophyll accumulation in the wild-type strain of C. reinhardtii. Interestingly, the increased level of carotenoids suggested that they might play a crucial role in cold stress acclimation. A temperature of 15 °C resulted in the highest carotenoid and chlorophyll productivity. At this temperature, carotenoid and chlorophyll productivity was 2 times and 1.3 times higher than at 25 °C, respectively. Subjecting a mutant defective in lutein and zeaxanthin accumulation to cold stress revealed that these two carotenoids are not essential for cold stress survival. Therefore, cold temperature could be used as a strategy to induce and increase the productivity of pigments in C. reinhardtii.

Sterols and squalene are essential biomolecules required for the homeostasis of eukaryotic membrane permeability and fluidity. Both compounds have beneficial effects on human health. As the current sources of sterols and squalene are plant and shark oils, microalgae are suggested as more sustainable sources. Nonetheless, the high costs of production and processing still hinder the commercialization of algal cultivation. Strain improvement for higher product yield and tolerance to harsh environments is an attractive way to reduce costs. Being an intermediate in sterol synthesis, squalene is converted to squalene epoxide by squalene epoxidase. This step is inhibited by terbinafine, a commonly used antifungal drug. In yeasts, some terbinafine-resistant strains overproduced sterols, but similar microalgae strains have not been reported. Mutants that exhibit greater tolerance to terbinafine might accumulate increased sterols and squalene content, along with the ability to tolerate the drug and other stresses, which are beneficial for outdoor cultivation. To explore this possibility, terbinafine-resistant mutants were isolated in the model green microalga Chlamydomonas reinhardtii using UV mutagenesis. Three mutants were identified and all of them exhibited approximately 50 percent overproduction of sterols. Under terbinafine treatment, one of the mutants also accumulated around 50 percent higher levels of squalene. The higher accumulation of pigments and triacylglycerol were also observed. Along with resistance to terbinafine, this mutant also exhibited higher resistance to oxidative stress. Altogether, resistance to terbinafine can be used to screen for strains with increased levels of sterols or squalene in green microalgae without growth compromise.

Biodiesel and natural products derived from microalgae require a smaller land area and have higher production rates compared to plants and animals and has recently attracted considerable interest. However, biodiesel production from microalgal triacylglycerol is still far from commercial realization due to its high production cost. One way to overcome this obstacle is to improve the triacylglycerol accumulation and couple its production with other high-value compounds. Of particular interest is the sterol biosynthetic pathway with squalene as an intermediate due to its close relationship with triacylglycerol and carotenoid biosynthetic pathways. Besides, both squalene and carotenoids are isoprenoid lipids that have health benefits. Perturbation of one pathway has been suggested to affect other pathways. Three terbinafine-sensitive mutants of the green microalga Chlamydomonas reinhardtii were isolated using terbinafine, a drug that inhibits squalene epoxidase, leading to squalene accumulation. One of the mutants, tfs2, accumulated twice the amount of wild-type triacylglycerol. As well as squalene accumulation, the presence of terbinafine further increased the triacylglycerol content. The level of prenyl lipid carotenoid and chlorophyll was also more significant than that of the wild type. Growth and photosynthesis were not compromised in this mutant. This is the first study that has demonstrated a mutant screening method to improve the co-production of TAG and isoprenoid lipids in a green microalgae.

Procyanidins, which are polyphenol compounds in grape seeds, apples, and berries, are known for their anti-inflammatory and antioxidant properties. In this study, we investigated durian flowers, an agricultural waste, as a novel procyanidin source. Durian trees bloom prolifically, but not all flowers develop into mature fruits, representing underutilized resources. Dual-platform metabolomic analysis using ultra-high-performance liquid chromatography with electrospray ionization quadrupole time-of-flight mass spectrometry and gas chromatography–mass spectrometry annotates polyphenols such as (−)-epicatechin, procyanidins B1, B2, and C1. The 80 % (v/v) ethanol extraction yielded a crude extract with a total procyanidin content of 7.68 mg/g. Bioactivity assays revealed that the procyanidin-rich crude extract reduced oxidative stress and exhibited anti-inflammatory effects against UVA in human keratinocytes (HaCaT). This study is the first to propose durian flowers as a sustainable and cost-effective procyanidin source with potential application in the nutraceutical and cosmeceutical industries, contributing significantly to the repurposing of agricultural waste through green technology.