Therapeutic potential of Lactobacillus casei and Chlorella vulgaris in high-fat diet-induced non-alcoholic fatty liver disease (NAFLD)-associated kidney damages: a stereological study.

BACKGROUND: The non-alcoholic fatty liver disease (NAFLD) is prevalent in as many as 25% of adults who are afflicted with metabolic syndrome. Oxidative stress plays a significant role in the pathophysiology of hepatic and renal injury associated with NAFLD. Therefore, probiotics such as Lactobacillus casei (LBC) and the microalga Chlorella vulgaris (CV) may be beneficial in alleviating kidney injury related to NAFLD. MATERIALS AND METHODS: This animal study utilized 30 C57BL/6 mice, which were evenly distributed into five groups: the control group, the NAFLD group, the NAFLD + CV group, the NAFLD + LBC group, and the NAFLD + CV + LBC group. A high-fat diet (HFD) was administered to induce NAFLD for six weeks. The treatments with CV and LBC were continued for an additional 35 days. Biochemical parameters, total antioxidant capacity (TAC), and the expression of kidney damage marker genes (KIM 1 and NGAL) in serum and kidney tissue were determined, respectively. A stereological analysis was conducted to observe the structural changes in kidney tissues. RESULTS: A liver histopathological examination confirmed the successful induction of NAFLD. Biochemical investigations revealed that the NAFLD group exhibited increased ALT and AST levels, significantly reduced in the therapy groups (p < 0.001). The gene expression levels of KIM-1 and NGAL were elevated in NAFLD but were significantly reduced by CV and LBC therapies (p < 0.001). Stereological examinations revealed reduced kidney size, volume, and tissue composition in the NAFLD group, with significant improvements observed in the treated groups (p < 0.001). CONCLUSION: This study highlights the potential therapeutic efficacy of C. vulgaris and L. casei in mitigating kidney damage caused by NAFLD. These findings provide valuable insights for developing novel treatment approaches for managing NAFLD and its associated complications.

Chlorella vulgaris algae ameliorates chlorpyrifos toxicity in Nile tilapia with special reference to antioxidant enzymes and Streptococcus agalactiae infection.

BACKGROUND: Chlorpyrifos (CPF) is a widely used pesticide in the production of plant crops. Despite rapid CPF biodegradation, fish were exposed to wastewater containing detectable residues. Recently, medicinal plants and algae were intensively used in aquaculture to replace antibiotics and ameliorate stress impacts. METHODS AND RESULTS: An indoor experiment was conducted to evaluate the deleterious impacts of CPF pollution on Nile tilapia health and the potential mitigation role of Chlorella vulgaris algae. Firstly, the median lethal concentration LC50 - 72 h of CPF was determined to be 85.8 µg /L in Nile tilapia (35.6 ± 0.5 g body weight) at a water temperature of 27.5 °C. Secondly, fish were exposed to 10% of LC50 - 72 h for six weeks, and tissue samples were collected and examined every two weeks. Also, Nile tilapia were experimentally infected with Streptococcus agalactiae. Exposed fish were immunosuppressed expressed with a decrease in gene expressions of interleukin (IL) 1ß, IL-10, and tumor necrosis factor (TNF)-α. Also, a decline was recorded in glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT) gene expression in the head kidney tissue. A high mortality rate (MR) of 100% was recorded in fish exposed to CPF for six weeks and challenged with S. agalactiae. Fish that received dietary C. vulgaris could restore gene expression cytokines and antioxidants compared to the control. After six weeks of CPF exposure, fish suffered from anemia as red blood cell count (RBCs), hemoglobin (Hb), and packed cell volume (PCV) significantly declined along with downregulation of serum total protein (TP), globulin (GLO), and albumin (ALB). Liver enzymes were significantly upregulated in fish exposed to CPF pollution, alanine aminotransferase (ALT) (42.5, 53.3, and 61.7 IU/L) and aspartate aminotransferase (AST) (30.1, 31.2, and 22.8) after 2, 4, and 6 weeks, respectively. On S. agalactiae challenge, high MR was recorded in Nile tilapia exposed to CPF (G3) 60%, 60%, and 100% in week 2, week 4, and week 6, and C. vulgaris provided a relative protection level (RPL) of 0, 14.29, and 20%, respectively. CONCLUSIONS: It was concluded that CPF pollution induces immunosuppressed status, oxidative stress, and anemic signs in Nile tilapia. In contrast, C. vulgaris at a 50 g/kg fish feed dose could partially ameliorate such withdrawals, restoring normal physiological parameters.

Microcystin-LR (MC-LR) inhibits green algae growth by regulating antioxidant and photosynthetic systems.

Microcystins release from bloom-forming cyanobacteria is considered a way to gain competitive advantage in Microcystis populations, which threaten water resources security and aquatic ecological balance. However, the effects of microcystins on microalgae are still largely unclear. Through simulated culture experiments and the use of UHPLC-MS-based metabolomics, the effects of two microcystin-LR (MC-LR) concentrations (400 and 1,600 µg/L) on the growth and antioxidant properties of three algae species, the toxic Microcystis aeruginosa, a non-toxic Microcystis sp., and Chlorella vulgaris, were studied. The MC-LR caused damage to the photosynthetic system and activated the protective mechanism of the photosynthetic system by decreasing the chlorophyll-a and carotenoid concentrations. Microcystins triggered oxidative stress in C. vulgaris, which was the most sensitive algae species studied, and secreted more glycolipids into the extracellular compartment, thereby destroying its cell structure. However, C. vulgaris eliminated reactive oxygen species (ROS) by secreting terpenoids, thereby resisting oxidative stress. In addition, two metabolic pathways, the vitamin B6 and the sphingolipid pathways, of C. vulgaris were significantly disturbed by microcystins, contributing to cell membrane and mitochondrial damage. Thus, both the low (400 µg/L) and the high (1,600 µg/L) MC-LR concentration inhibited algae growth within 3 to 7 days, and the inhibition rates increased with the increase in the MC-LR concentration. The above results indicate that the toxin-producing Microcystis species have a stronger toxin tolerance under longer-term toxin exposure in natural water environments. Thus, microcystins participates in interspecific interaction and phytoplankton population regulation and creates suitable conditions for the toxin-producing M. aeruginosa to become the dominant species in algae blooms.

Impact of organic matter molecular weight on hexavalent chromium enrichment in green microalgae.

In lightly polluted water containing heavy metals, organic matter, and green microalgae, the molecular weight of organic matter may influence both the growth of green microalgae and the concentration of heavy metals. This study elucidates the effects and mechanisms by which different molecular weight fractions of fulvic acid (FA), a model dissolved organic matter component, facilitate the bioaccumulation of hexavalent chromium (Cr(VI)) in a typical green alga, Chlorella vulgaris. Findings show that the addition of FA fractions with molecular weights greater than 10 kDa significantly enhances the enrichment of total chromium and Cr(VI) in algal cells, reaching 21.58%-31.09 % and 16.17 %-22.63 %, respectively. Conversely, the efficiency of chromium enrichment in algal cells was found to decrease with decreasing molecular weight of FA. FA molecular weight within the range of 0.22 µm-30 kDa facilitated chromium enrichment primarily through the algal organic matter (AOM) pathway, with minor contributions from the algal cell proliferation and extracellular polymeric substances (EPS) pathways. However, with decreasing FA molecular weight, the AOM and EPS pathways become less prominent, whereas the algal cell proliferation pathway becomes dominant. These findings provide new insights into the mechanism of chromium enrichment in green algae enhanced by medium molecular weight FA.

Predicting the Structure of Enzymes with Metal Cofactors: The Example of [FeFe] Hydrogenases.

The advent of deep learning algorithms for protein folding opened a new era in the ability of predicting and optimizing the function of proteins once the sequence is known. The task is more intricate when cofactors like metal ions or small ligands are essential to functioning. In this case, the combined use of traditional simulation methods based on interatomic force fields and deep learning predictions is mandatory. We use the example of [FeFe] hydrogenases, enzymes of unicellular algae promising for biotechnology applications to illustrate this situation. [FeFe] hydrogenase is an iron-sulfur protein that catalyzes the chemical reduction of protons dissolved in liquid water into molecular hydrogen as a gas. Hydrogen production efficiency and cell sensitivity to dioxygen are important parameters to optimize the industrial applications of biological hydrogen production. Both parameters are related to the organization of iron-sulfur clusters within protein domains. In this work, we propose possible three-dimensional structures of Chlorella vulgaris 211/11P [FeFe] hydrogenase, the sequence of which was extracted from the recently published genome of the given strain. Initial structural models are built using: (i) the deep learning algorithm AlphaFold; (ii) the homology modeling server SwissModel; (iii) a manual construction based on the best known bacterial crystal structure. Missing iron-sulfur clusters are included and microsecond-long molecular dynamics of initial structures embedded into the water solution environment were performed. Multiple-walkers metadynamics was also used to enhance the sampling of structures encompassing both functional and non-functional organizations of iron-sulfur clusters. The resulting structural model provided by deep learning is consistent with functional [FeFe] hydrogenase characterized by peculiar interactions between cofactors and the protein matrix.

Inorganic carbon limitation decreases ammonium removal and N2O production in the algae-nitrifying bacteria symbiosis system.

Ammonium removal by a symbiosis system of algae (Chlorella vulgaris) and nitrifying bacteria was evaluated in a long-term photo-sequencing batch reactor under varying influent inorganic carbon (IC) concentrations (15, 10, 5 and 2.5 mmol L-1) and different nitrogen loading rate (NLR) conditions (270 and 540 mg-N L-1 d-1). The IC/N ratios provided were 2.33, 1.56, 0.78 and 0.39, respectively, for an influent NH4+-N concentration of 90 mg-N L-1 (6.43 mmol L-1). The results confirmed that both ammonium removal and N2O production were positively related with IC concentration. Satisfactory ammonium removal efficiencies (>98 %) and rates (29-34 mg-N gVSS-1 h-1) were achieved regardless of NLR levels under sufficient IC of 10 and 15 mmol L-1, while insufficient IC at 2.5 mmol L-1 led to the lowest ammonium removal rates of 0 mg-N gVSS-1 h-1. The ammonia oxidation process by ammonia oxidizing bacteria (AOB) played a predominant role over the algae assimilation process in ammonium removal. Long-time IC deficiency also resulted in the decrease in biomass and pigments of algae and nitrifying bacteria. IC limitation led to the decreasing N2O production, probably due to its negative effect on ammonia oxidation by AOB. The optimal IC concentration was determined to be 10 mmol L-1 (i.e., IC/N of 1.56, alkalinity of 500 mg CaCO3 L-1) in the algae-bacteria symbiosis reactor, corresponding to higher ammonia oxidation rate of ∼41 mg-N gVSS-1 h-1 and lower N2O emission factor of 0.13 %. This suggests regulating IC concentrations to achieve high ammonium removal and low carbon emission simultaneously in the algae-bacteria symbiosis wastewater treatment process.

Removal of polycyclic aromatic hydrocarbons (PAHs) from produced water using the microalgae Chlorella vulgaris cultivated in mixotrophic and heterotrophic conditions.

Chlorella vulgaris was cultivated for 15 days in 10 different treatments under mixotrophic and heterotrophic conditions, using wastewater from oil and poultry industries as the culture medium. The blends were made with produced water (PW), sterilized produced water (PWs), sterilized poultry wastewater (PoWs), sterilized seawater (SWs), and the addition of sodium nitrate to evaluate cell growth in treatments and the removal of PAHs. The heterotrophic condition showed more effective removal, having an initial concentration of 3.93 µg L-1 and a final concentration of 0.57 µg L-1 of total PAHs reporting 83%, during phycoremediation of (PW) than the mixotrophic condition, with an initial concentration of 3.93 µg L-1 and a final concentration of 1.96 and 43% removal for the PAHs. In the heterotrophic condition, the blend with (PWs + SWs) with an initial concentration of 0.90 µg L-1 and a final concentration of 0.32 µg L-1 had 64% removal of total PAHs compared to the mixotrophic condition with 37% removal having an initial concentration of 0.90 µg L-1 and a final concentration of 0.56 µg L-1. However, the best result in the mixotrophic condition was obtained using a blend of (PWs + PoWs) that had an initial cell concentration of 1.18 × 105 cells mL-1 and reached a final cell concentration of 4.39 × 105 cells mL-1, an initial concentration of 4.76 µg L-1 and a final concentration of 0.37 µg L-1 having a 92% total removal of PAHs. The biostimulation process increased the percentage of PAHs removal by 45% (PW) in the mixotrophic condition. This study showed that it is possible to allow an environmental remediation strategy that significantly reduces effluent toxicity and generates high value-added biomass in contaminated effluents rich in nutrients and carbon, based on a circular bioeconomy model.

Nutrients recovery from dairy wastewater by Chlorella vulgaris and comparison of the lipid's composition with various chlorella strains for biodiesel production.

Microalgae biomass is regarded as a promising feedstock for biodiesel production. The biomass lipid content and fatty acids composition are among the main selective criteria when screening microalgae strains for biodiesel production. In this study, three strains of Chlorella microalgae (C. kessleri, C. sorokiniana, C. vulgaris) were cultivated nutrient media with different nitrogen contents, and on a medium with the addition of dairy wastewater. Moreover, microalgae grown on dairy wastewater allowed the removal of azote and phosphorous. The removal efficiency of 90%, 53% and 95% of ammonium nitrogen, total nitrogen and phosphate ions, respectively, were reached. The efficiency of wastewater treatment from inorganic carbon was 55%, while the maximum growth of biomass was achieved. All four samples of microalgae had a similar fatty acid profile. Palmitic acid (C16:0) was the most abundant saturated fatty acid (SFA), and is suitable for the production of biodiesel. The main unsaturated fatty acids (UFA) present in the samples were oleic acid (C18:1 n9); linoleic acid (C18:2 n6) and alpha-linolenic acid (C18:3 n3), which belong to omega-9, omega-6, omega-3, respectively.

Presence of humic acid in the environment holds promise as a potential mitigating factor for the joint toxicity of polystyrene nanoplastics and herbicide atrazine to Chlorella vulgaris: 96-H acute toxicity.

Increasing amounts of amino-functionalized polystyrene nanoplastics (PS-NH2) are entering aquatic ecosystems, raising concerns. Hence, this study investigated 96-h acute toxicity of PS-NH2 and its combination with the pesticide atrazine (ATZ) in the absence/presence of humic acid (HA) on the microalgae Chlorella vulgaris (C. vulgaris). Results showed that both PS-NH2 and PS-NH2+ATZ reduced algal growth, photosynthetic pigments, protein content, and antioxidant capacity, while increasing enzymatic activities. Gene expression related to oxidative stress was altered in C. vulgaris exposed to these treatments. Morphological and intracellular changes were also observed. The combined toxicity of PS-NH2+ATZ demonstrated a synergistic effect, but the addition of environmentally relevant concentration of HA significantly alleviated its toxicity to C. vulgaris, indicating an antagonistic effect due to the emergence of an eco-corona, and entrapment and sedimentation of PS-NH2+ATZ particles by HA. This study firstly highlights the role of HA in mitigating the toxicity of PS-NH2 when combined with other harmful compounds, enhancing our understanding of HA's presence in the environment.

Prediction of acute toxicity for Chlorella vulgaris caused by tire wear particle-derived compounds using quantitative structure-activity relationship models.

Tire wear particles (TWPs) enter aquatic ecosystems through various pathways, such as rainwater and urban runoff. Additives in TWPs can harm aquatic organisms in these ecosystems. Therefore, it is essential to investigate their toxicity to aquatic organisms. In our study, we initially recorded the median effective concentrations of 21 TWP-derived compounds on Chlorella vulgaris growth, ranging from 0.04 to 8.60 mg/L. Subsequently, through an extensive review of the literature, we incorporated 112 compounds with specific toxicity endpoints to construct the QSAR model using genetic algorithm and multiple linear regression techniques, followed by the construction of the consensus model and the quantitative read-across structure-activity relationship (q-RASAR) model. Meanwhile, we employed rigorous internal and external validation measures to assess the performance of the model. The results indicated that the developed q-RASAR model exhibited strong adaptation, robustness, and reliable prediction, with q-RASAR indicators of Q2LOO = 0.7673, R2tr = 0.8079, R2test = 0.8610, Q2Fn = 0.8285-0.8614, and CCCtest = 0.9222. Based on an external dataset containing 128 emerging TWP-derived compounds, the model's applicability domain coverage was 90.6 %. The q-RASAR model predicted that the structure of diphenylamine was associated with higher toxicity, possibly liked to the SpMax2_Bhm and LogBCF descriptors. The established model reliably provides prediction and fills a critical data gap. These findings highlight the potential risks posed by emerging TWP-derived compounds to aquatic organisms.

Molecular mechanism of dissolvable metal nanoparticles-enhanced CO2 fixation by algae: Metal-chlorophyll synthesis.

Algae-driven photosynthetic CO2 fixation is a promising strategy to mitigate global climate changes and energy crises. Yet, the presence of metal nanoparticles (NPs), particularly dissolvable NPs, in aquatic ecosystems introduces new complexities due to their tendency to release metal ions that may perturb metabolic processes related to algal CO2 fixation. This study selected six representative metal NPs (Fe3O4, ZnO, CuO, NiO, MgO, and Ag) to investigate their impacts on CO2 fixation by algae (Chlorella vulgaris). We discovered an intriguing phenomenon that bivalent metal ions released from the metal NPs, especially from ZnO NPs, substituted Mg2+ within the porphyrin ring. This interaction led to 81.8% and 76.1% increases in Zinc-chlorophyll and Magnesium-chlorophyll contents within algal cells at 0.01 mM ZnO NPs, respectively. Integrating metabolomics and transcriptomics analyses revealed that ZnO NPs mainly promoted the photosynthesis-antenna protein pathway, porphyrin and chlorophyll metabolism, and carbon fixation pathway, thereby mitigating the adverse effects of Zn2+ substitution in light harvesting and energy transfer for CO2 fixation. Ultimately, the genes encoding Rubisco large subunit (rbcL) responsible for CO2 fixation were upregulated to 2.60-fold, resulting in a 76.3% increase in carbon fixation capacity. Similar upregulations of rbcL expression (1.13-fold) and carbon fixation capacity (76.1%) were observed in algal cells even at 0.001 mM ZnO NPs, accompanied by valuable lipid accumulation. This study offers novel insights into the molecular mechanism underlying NPs on CO2 fixation by algae and potentially introduces strategies for global carbon sequestration.

Modeling and optimization of biomass productivity of Chlorella vulgaris using response surface methodology, analysis of variance and machine learning for carbon dioxide capture.

This study explores bioremediation's effectiveness in reducing carbon emissions through the use of microalgae Chlorella vulgaris, known for capturing carbon dioxide and producing biomass. The impact of temperature and light intensity on productivity and carbon dioxide capture was investigated, and cultivation conditions were optimized in a photobioreactor using response surface methodology (RSM), analysis of variance (ANOVA), and deep neural networks (DNN). The optimal conditions determined were 28.74 °C and 225 µmol/m2/s with RSM, and 29.55 °C and 226.77 µmol/m2/s with DNN, closely aligning with literature values (29 °C and 225 µmol/m2/s). DNN demonstrated superior performance compared to RSM, achieving higher accuracy due to its capacity to process larger datasets using epochs and batches. The research serves as a foundation to further in this field by demonstrating the potential of utilizing diverse mathematical models to optimize bioremediation conditions, and offering valuable insights to improve carbon dioxide capture efficiency in microalgae cultivation.

Low-cost pyrolysis of biomass-derived nitrogen-doped porous carbon: Chlorella vulgaris replaces melamine as a nitrogen source.

Porous carbon generated from biomass has a rich pore structure, is inexpensive, and has a lot of promise for use as a carbon material for energy storage devices. In this work, nitrogen-doped porous carbon was prepared by co-pyrolysis using bagasse as the precursor and chlorella as the nitrogen source. ZnCl2 acts as both an activator and a nitrogen fixer during activation to generate pores and reduce nitrogen loss. The thermal weight loss experiments showed that the pyrolysis temperatures of bagasse and chlorella overlap, which created the possibility for the synthesis of nitrogen-rich biochar. The optimum sample (ZBC@C-5) possessed a surface area of 1508 m2g-1 with abundant nitrogen-containing functional groups. ZBC@C-5 in the three-electrode system exhibited 244.1F/g at 0.5A/g, which was extremely close to ZBC@M made with melamine as the nitrogen source. This provides new opportunities for the use of low-cost nitrogen sources. Furthermore, the devices exhibit better voltage retention (39%) and capacitance retention (96.3%). The goal of this research is to find a low cost, and effective method for creating nitrogen-doped porous carbon materials with better electrochemical performance for highly valuable applications using bagasse and chlorella.

Combined toxic effects of polystyrene nanoplastics and lead on Chlorella vulgaris growth, membrane lipid peroxidation, antioxidant capacity, and morphological alterations.

In recent years, there has been a significant rise in the utilization of amino-functionalized polystyrene nanoplastics (PS-NH2). This surge in usage can be attributed to their exceptional characteristics, including a substantial specific surface area, high energy, and strong reactivity. These properties make them highly suitable for a wide range of industrial and medical applications. Nevertheless, there is a growing apprehension regarding their potential toxicity to aquatic organisms, particularly when considering the potential impact of heavy metals like lead (Pb) on the toxicity of PS-NH2. Herein, we examined the toxic effects of sole PS-NH2 (90 nm) at five concentrations (e.g., 0, 0.125, 0.25, 0.5, and 1 mg/L), as well as the simultaneous exposure of PS-NH2 and Pb2+ (using two environmental concentrations, e.g., 20 µg/L for Pb low (PbL) and 80 µg/L for Pb higher (PbH)) to the microalga Chlorella vulgaris. After a 96-h exposure, significant differences in chlorophyll a content and algal growth (biomass) were observed between the control group and other treatments (ANOVA, p < 0.05). The algae exposed to PS-NH2, PS-NH2 + PbL, and PS-NH2 + PbH treatment groups exhibited dose-dependent toxicity responses to chlorophyll a content and biomass. According to the Abbott toxicity model, the combined toxicity of treatment groups of PS-NH2 and PbL,H showed synergistic effects. The largest morphological changes such as C. vulgaris' size reduction and cellular aggregation were evident in the medium treated with elevated concentrations of both PS-NH2 and Pb2+. The toxicity of the treatment groups followed the sequence PS-NH2 < PS-NH2 + PbL < PS-NH2 + PbH. These results contribute novel insights into co-exposure toxicity to PS-NH2 and Pb2+ in algae communities.

Photoheterotroph improved the growth and nutrient levels of Chlorella vulgaris and the related molecular mechanism.

Microalgae are rich in fatty acids, proteins, and other nutrients, which have gained the general attention of researchers all over the world. For the development of Chlorella vulgaris in food and feed industry, this study was conducted to investigate the differences in C. vulgaris' growth and nutritional components under different culture conditions (autotrophic, heterotrophic, photoheterotrophic) and the internal factors through cell counting in combination with transcriptome and nutrient analyses. The results showed that, under the photoheterotrophic condition, Chlorella's growth and the contents of lipid and protein were significantly higher than that under the heterotrophic condition, and the moisture content was lower than that under the heterotrophic condition. The saturated fatty acid content under the photoheterotrophic condition was the lowest, while the polyunsaturated fatty acid content was significantly higher than those under the other two conditions. There were 46,583 differentially expressed genes (DEGs), including 33,039 up-regulated DEGs (70.93%) and 13,544 down-regulated DEGs (29.07%), under the photoheterotrophic condition in comparison with the autotrophic condition. The fold change between the two conditions of samples of up-regulated genes was higher than that of the down-regulated genes. The KEGG enrichment showed that the up-regulated DEGs in the photoheterotrophic condition were significantly enriched in 5 pathways, including protein processing in endoplasmic reticulum pathway, photosynthesis pathway, photosynthesis-antenna protein pathway, endocytosis pathway, and phosphonate and phosphinate metabolism pathway. DEGs related to fatty acid metabolic pathways were significantly enriched in the fatty acid biosynthesis pathway and the biosynthesis of unsaturated fatty acid pathway. The qPCR analysis showed that the expression pattern of the selected genes was consistent with that of transcriptome analysis. The results of this study lay a theoretical foundation for the large-scale production of Chlorella and its application in food, feed, and biodiesel. KEY POINTS: • Nutrient levels under photoheterotrophic condition were higher than other conditions. • Six important pathways were discovered that affect changes in nutritional composition. • Explored genes encode important enzymes in the differential metabolic pathways.

Innovative vs classical methods for drying heterotrophic Chlorella vulgaris: Impact on protein quality and sensory properties.

Drying is a necessary step in the microalgae production chain to reduce microbial load and oxidative degradation of the end product. Depending on the differences in applied temperature and treatment time, the process of drying can have a substantial impact on protein quality and aroma, important characteristics determining the incorporation potential in food products. In this study, we compared the drying of heterotrophic Chorella vulgaris with both innovative (agitated thin film drying (ATFD), pulse combustion drying (PCD) and solar drying (SolD)) and commonly used drying techniques (spray drying (SprD) and freeze drying (FD)). To evaluate the impact on protein quality, we evaluated techno-functional properties, in vitro digestibility (INFOGEST) as well as protein denaturation using differential scanning calorimetry (DSC). A sensory analysis was performed by a trained expert panel, combined with headspace solid-phase microextraction (HS-SPME) - gas chromatography-mass spectrometry (GC-MS) to determine volatile organic compounds (VOCs). ATFD was found to increase techno-functional properties such as gelling, water holding and solubility as well as in vitro protein digestibility. These observations could be related to induced cell disruption and protein denaturation by ATFD. Sensory analysis indicated an increased earthy off-flavor after ATFD. Interestingly, the high-temperature PCD led to an increase in cacao odor while low-temperature FD resulted in lower flavor, odors and VOCs. These results demonstrate that protein quality and sensorial properties of C. vulgaris can be steered through the type of drying, which could help in the selection of application-specific drying methods. Overall, this work could promote the incorporation of microalgal single cell proteins in different innovative food products.

Pulsed electric field treatment for preservation of Chlorella suspensions and retention of gelling capacity.

Pulsed electric field (PEF) processing has emerged as an alternative to thermal pasteurization for the shelf-life extension of heat-sensitive liquids at industrial scale. It offers the advantage of minimal alteration in physicochemical characteristics and functional properties. In this study, a pilot-scale continuous PEF processing (Toutlet < 55 °C) was applied to microalgae Chlorella vulgaris (Cv) suspensions (pH = 6.5), which was proposed as a functional ingredient for plant-based foods. Cv suspensions were inoculated with three distinct food spoilage microorganisms (Pseudomonas guariconensis, Enterobacter soli and Lactococcus lactis), isolated from the Cv biomass. PEF treatments were applied with varying electric field strength Eel of 16 to 28 kV/cm, pulse repetition rate f of 100 to 140 Hz, with a pulse width τ of 20 µs and an inlet product temperature Tin of 30 °C. The aim was to evaluate the PEF-induced microbial reduction and monitor the microbial outgrowth during a 10-day cold storage period (10 °C). Maximum inactivation of 4.1, 3.7 and 3.6 logs was achieved (28 kV/cm and 120 Hz) for the investigated isolates, respectively. Under these conditions, the critical electric field strengths Ecrit, above which inactivation was observed, ranged from 22.6 to 24.6 kV/cm. Moreover, repeated PEF treatment resulted in similar inactivation efficiency, indicating its potential to enhance shelf-life further.

Enhancing microalgae biomass production: Exploring improved scraping frequency in a hybrid cultivation system.

Recently, hybrid systems, such as those incorporating high-rate algal ponds (HRAPs) and biofilm reactors (BRs), have shown promise in treating domestic wastewater while cultivating microalgae. In this context, the objective of the present study was to determine an improved scraping frequency to maximize microalgae biomass productivity in a mix of industrial (fruit-based juice production) and domestic wastewater. The mix was set to balance the carbon/nitrogen ratio. The scraping strategy involved maintaining 1 cm wide stripes to retain an inoculum in the reactor. Three scraping frequencies (2, 4, and 6 days) were evaluated. The findings indicate that a scraping frequency of each 2 days provided the highest biomass productivity (18.75 g total volatile solids m-2 d-1). The species' behavior varied with frequency: Chlorella vulgaris was abundant at 6-day intervals, whereas Tetradesmus obliquus favored shorter intervals. Biomass from more frequent scraping demonstrated a higher lipid content (15.45%). Extrapolymeric substance production was also highest at the 2-day frequency. Concerning wastewater treatment, the system removed 93% of dissolved organic carbon and ∼100% of ammoniacal nitrogen. Combining industrial and domestic wastewater sources to balance the carbon/nitrogen ratio enhanced treatment efficiency and biomass yield. This study highlights the potential of adjusting scraping frequencies in hybrid systems for improved wastewater treatment and microalgae production.

Taguchi approach for assessing supercritical CO2 (sCO2) fluid extraction of polyhydroxyalkanoate (PHA) from Chlorella Vulgaris sp. microalgae.

This research explicitly investigates the utilization of Chlorella Vulgaris sp. microalgae as a renewable source for lipid production, focusing on its application in bioplastic manufacturing. This study employed the supercritical fluid extraction technique employing supercritical CO2 (sCO2) as a green technology to selectively extract and produce PHA's precursor utilizing CO2 solvent as a cleaner solvent compared to conventional extraction method. The study assessed the effects of three extraction parameters, namely temperature (40-60 °C), pressure (15-35 MPa), and solvent flow rate (4-8 ml/min). The pressure, flowrate, and temperature were found to be the most significant parameters affecting the sCO2 extraction. Through Taguchi optimization, the optimal parameters were determined as 60 °C, 35 MPa, and 4 ml/min with the highest lipid yield of 46.74 wt%; above-average findings were reported. Furthermore, the pretreatment process involved significant effects such as crumpled and exhaustive structure, facilitating the efficient extraction of total lipids from the microalgae matrix. This study investigated the microstructure of microalgae biomatrix before and after extraction using scanning electron microscopy (SEM) and thermogravimetric analysis (TGA). Fourier-transform infrared spectroscopy (FTIR) was utilized to assess the potential of the extracted material as a precursor for biodegradable plastic production, with a focus on reduced heavy metal content through inductively coupled plasma-optical emission spectrometry (ICP-OES) analysis. The lipid extracted from Chlorella Vulgaris sp. microalgae was analysed using gas chromatography-mass spectrometry (GC-MS), identifying key constituents, including oleic acid (C18H34O2), n-Hexadecanoic acid (C16H32O2), and octadecanoic acid (C18H36O2), essential for polyhydroxyalkanoate (PHA) formation.

Chlorella alleviates the intestinal damage of tilapia caused by microplastics.

Polyethylene microplastics (MPs) of the different sizes may result in different response in fish. Studies showed microorganisms adhered to the surface of MPs have toxicological effect. Juveniles tilapia (Oreochromis niloticus, n = 600, 26.5 ± 0.6 g) were dispersed into six groups: the control group (A), 75 nm MP exposed group (B), 7.5 µm group (C) and 750 (D) µm group, 75 nm + 7.5 µm+750 µm group (E) and 75 nm + Chlorella vulgaris group (F), and exposed for 10 and 14 days. The intestinal histopathological change, enzymic activities, and the integrated "omics" workflows containing transcriptomics, proteomics, microbiota and metabolomes, have been performed in tilapia. Results showed that MPs were distributed on the surface of goblet cells, Chlorella group had severe villi fusion without something like intestinal damage, as in other MPs groups. The intestinal Total Cholesterol (TC, together with group E) and Tumor Necrosis Factor α (TNFα, except for group B) contents in group F were significantly increased, cytochrome p450 1a1 (EROD, group B and E) significantly increased, adenosine triphosphate (ATP), lipoprotein lipase (LPL) and caspase 3 (except group B) also significantly increased at 14 d. At 14 days, group E saw considerably higher regulation of the actin cytoskeleton, focal adhesion, insulin signaling pathway, and AGE-RAGE signaling pathway in diabetes complications. Whereas, chlorella enhanced the focal adhesion, cytokine-cytokine receptor interaction, and MAPK signaling pathways. PPAR signaling pathway has been extremely significantly enriched via the proteomics method. Candidatus latescibacteria, C. uhrbacteria, C. abyssubacteria, C. cryosericota significantly decreased caused by MPs of different particle sizes. Carboxylic acids and derivatives, indoles and derivatives, organooxygen compounds, fatty acyls and organooxygen compounds significantly increased with long-term duration, especially PPAR signaling pathway. MPs had a size-dependent long-term effect on histopathological change, gene and protein expression, and gut microbial metabolites, while chlorella alleviates the intestinal histopathological damage via the integrated "omics" workflows.