Investigation of anaerobic digestion of Chlorella sp. and Micractinium sp. grown in high-nitrogen wastewater and their co-digestion with waste activated sludge

This study investigated two wildtype green algae, Micractinium sp. and Chlorella sp., for their growth in high nitrogen wastewater (mixture of sludge centrate and primary effluent wastewater) and subsequent anaerobic digestion under mesophilic conditions. Extraction and analysis of extracellular polymeric substances (EPS) in both algal species during cultivation showed that Micractinium generated larger quantity of EPS-proteins than Chlorella. Anaerobic digestion of harvested algae showed the opposite trend that Chlorella allowed a higher CH₄ yield on the volatile solids fed the digester (VS_fed) of 230 dm³ kg⁻¹ than Micractinium (209 dm³ kg⁻¹). These results suggested that different growth patterns of two types of algae, with different quantity of EPS expressed, affected anaerobic digestibility and biogas yield. Co-digestion of algae with waste activated sludge (WAS) improved the volatile solids reduction, hydrolysis efficiency as well as the biogas yields of algae.     

Photoautotrophic hydrogen production by nitrogen-deprived Chlamydomonas reinhardtii cultures

In this study we have demonstrated the possibility of phototrophic hydrogen production in C. reinhardtii under N-deprived conditions. When tested under air + CO₂, and Ar + CO₂ N-deprived C. reinhardtii demonstrated decrease in PSII activity mainly due to over reduction of PQ, in addition no ascorbate accumulation was observed in cells. Under air + CO₂ atmosphere cells accumulated excessive amounts of starch. When incubated under Ar + CO₂ atmosphere cells accumulated starch as nitrogen replete cultures and no hydrogen production was observed. Hydrogen production (86 ml H₂ per one l of culture) occurred under Ar + CO₂ atmosphere when particular two-step illumination protocol was implicated. In oxygen producing and early oxygen consuming stage cells were illuminated under light intensity 169 μE m^?2 s^?1. When light was switched to 30 μE m^?2 s^?1, cultures quickly respired all oxygen and transient to anaerobic conditions with subsequent hydrogen production 2 h later. Actual quantum yield of C. reinhardtii cultures was measured in photobioreactor and maximal quantum efficiency of PSII of dark adapted cells together with JIP test were studied.     

Very efficient dehydrogenation of methanolysis reaction with nitrogen doped Chlorella Vulgaris microalgae carbon as metal-free catalysts

In this study, nitrogen (N) doped metal-free catalysts were obtained as a result of nitric acid (HNO₃) activation of carbon sample (C–KOH–N), which was obtained based on Chlorella Vulgaris microalgae by KOH activation (C–KOH). These catalysts have been effectively used to produce hydrogen (H₂) from the sodium borohydride (NaBH₄) methanolysis reaction. Compared to the C–KOH catalyst, the catalytic activity for C–KOH–N showed a seven-fold improvement. Hydrogen generation rate (HGR) values obtained for the NaBH₄ methanolysis reaction for C–KOH and C–KOH–N metal-free catalysts were 3250 and 20,100 mL min^?1 g^?1. The catalysts were characterized using various analytical techniques such as XPS, XRD, SEM, FTIR, BET, and elemental analysis. This work can provide a new alternative strategy to produce specific heteroatom-doped metal-free carbon catalysts for environmentally friendly conversion to produce H₂ efficiently.     

Photosynthetic microbial desalination cell (PhMDC) using Chlamydomonas sp. (UKM6) and Scenedesmus sp. (UKM9) as biocatalysts for electricity production and water treatment

A current Microbial Desalination Cell (MDC) system often uses chemicals or air cathodes that are toxic and impractical for scalable applications. This work presents the MDC performance that utilized microalgae species Chlamydomonas sp. (UKM6) and Scenedesmus sp. (UKM9). These species supported the role of the terminal electron acceptor in the cathode chamber of photosynthetic MDCs (PhMDC). The results showed that PhMDC-UKM9 and UKM6 produced 1942 mW/m³ and 1714 mW/m³ power densities with 44% and 32% desalination rates, respectively. The desalination of salt concentration (35 g/L) was approaching the practical application of seawater. Both UKM6 and UKM9 achieved chemical oxygen demand (COD) removal in the anode chamber by 49% and 53%, respectively. 16S microbial community analysis in anolyte revealed phylum Firmicutes as the dominion community. This study demonstrated that the local microalgae species integrate power production, wastewater treatment, and water purification through PhMDC operation, comparable with other studies using commercial microalgae.     

Enhancement of lipid productivity in green microalgae Chlorella sp. via fast neutron irradiation

Microalgae is becoming a promising candidate for biofuel production and energetic lipid production, which urges the need to develop efficient methods for improvement of the lipid production. In this study, the neutron irradiation was employed to improve lipid production under different dosage irradiation. Effects of different dosages (No.1:8.14 × 10⁹, No.2:5.64 × 10⁸, No.4:9.42 × 10⁷ n/cm²) were compared by investigating associated algae cell growth, total lipid production, metabolization and enzyme activity. Dosages No.2 and No.4 improved the lipid content without negative influence on the growing, whereas irradiations shortened the period of biomass accumulation. The neutron irradiation in No.2 accelerated the synthesis of lipid from the carbohydrate transition process. In addition, the neutron irradiation not only increased the total lipid production by 20% but also speeded up the reaction rate to reach the maximal total lipid production in 28 days. Our results can provide a better understanding of the lipid production in microalgae Chlorella sp. by neutron irradiation, which are critical for evaluation of neutron irradiation based technologies for the sustainable and renewable biofuel production.     

Biomass harvesting and concentration of microalgae scenedesmus sp. cultivated in a pilot phobioreactor

Samples of a culture of Scenedesmus sp. grown in a pilot raceway photo bioreactor were processed in the laboratory by flocculation and centrifugation, to assess the efficiency of solid concentration of both methods. Three flocculation agents were tested, ferrous sulfate (FeSO₄), ferric chloride (FeCl₃) and aluminum sulfate Al₂(SO4)₃ using concentrations of 0.05, 0.2, 0.8, 1.5, 3.0 y 7.0 g L⁻¹ and pH range between 4.0 and 11.0 for the better concentration response of each flocculant. Additional flocculation tests were carried out to measure the sedimentation kinetics of the concentrated biomass. All these tests were performed with culture samples of 800 cm³ for 12 min of stirring and 10 min of sedimentation with three repetitions. In the case of centrifugation the testing was carried out using a laboratory centrifuge run at speeds of 1500, 1800 and 2200 rpm with culture samples of 300 cm³ for 15 min and three repetitions. In order to quantify the efficiency of the concentration, initial and final turbidity of the cleared water and concentrated portion were measured. The flocculation experiments showed that a limit of maximum concentration efficiency of 97.9% that was reached with 1.5 g L⁻¹ of Al₂(SO₄)₃ at pH of 8.5 and an average sedimentation velocity of about 2.7 cm min⁻¹. Testing also showed that FeSO₄ was the worst flocculant agent in the range tested. With respect to the centrifugation test, the solid concentration efficiency varied from 95.2% at speed of 1500 rpm up to 96.0% at speed of 2200 rpm.     

Growth and hydrogen production by three Chlamydomonas strains cultivated in a commercial fertilizer

In the present investigation, we report the growth and hydrogen production of two wild type Chlamydomonas reinhardtii strains isolated from a tropical oxidation pond in Costa Rica. The performance of these two new isolates was compared to that of Chlamydomonas reinhardtii CC124. All the strains were grown both in conventional Tris-Acetate-Phosphate medium (TAP) and in a commercial fertilizer medium (NPK 20-20-20). The growth of the new two isolates in medium formulated with fertilizer was higher than that attained with the reference strain (CC124). However, the hydrogen production performance of the strain CC124 in TAP-S and fertilizer were comparable, while the two new strains performed better in fertilizer, although the total hydrogen production was lower than that achieved with CC124. By using fertilizer it is possible to reduce the cost of chemical reagents by about 63% compared to TAP. Another advantage of the fertilizer is that it does not contain sulfur, therefore it can be directly used for hydrogen production using the Melis protocol.     

光温培养条件对小球藻Chlorella sp.生长及产物的影响

光温条件对微藻培养具有重要作用。采用小球藻Chlorella sp.进行了不同光照强度、光照时间和温度的培养试验,研究了光温条件对小球藻生长速率、生物量、生物组分及脂肪酸组成的影响。研究结果表明:光照强度对小球藻的最终生物量影响不大,但对生长速率、日产量和生物组分影响明显,蛋白质和脂肪酸含量随着光照强度的升高而减少,糖类物质随着光照强度的增加而增加;光照时间对小球藻糖类的累积产生较大影响,对脂肪酸的累积影响相对不太明显,对脂肪酸的组成产生一定的影响;温度对小球藻的生长速率、生物量、油脂含量和油脂产量都有显著的影响。因此,在藻类的培养过程中,适当地调节光温条件有利于目标产物的生产。     

Growth promotion of Chlorella vulgaris by modification of nitrogen source composition with symbiotic bacteria,Microbacterium sp. HJ1

To increase algal growth in treated livestock waste water, we designed a culture system targeting symbiotic bacteria. Microbacterium sp. HJ1 is a symbiotic bacteria associated with Chlorella vulgaris, which was found to increase the growth rate when controlled by nitrogen addition. The validated analysis model for nitrogen source mixture was used to analyze the growth and final pH of Microbacterium sp. HJ1, in different compositions of nitrogen sources, by elucidating the functions of each nitrogen ions such as NO₃⁻, NO₂⁻, and NH₄⁺. By modifying the growth medium made from treated livestock waste water with additional nitrogen source, we were able to increase dry cell weight (DCW) of C. vulgaris by 65.7% and chlorophyll a contents by 78.8%. This is an example of an indirect method to increase algal biomass by changing the population of symbiotic bacteria, and it is the practical application of positive effects from symbiotic bacteria to the host.     

Cultivating the marine microalga Nannochloropsis salina under various nitrogen sources: Effect on biovolume yields,lipid content and composition,and invasive organisms

Algae can be a viable source for biofuel production, but the source of nitrogen used to cultivate could affect algae yields. Here, we observe how various nitrogen treatments can impact the growth and biovolume of microalga Nannochloropsis salina as well as invasion of undesired organisms. Invading organisms increase the likelihood of crashes of the desired microalgae culture. Experiments were conducted over 28 days in open aquaria in a greenhouse. We used five different nitrogen treatments; ammonium chloride (NH₄Cl), ammonium hydroxide (NH₄OH), sodium nitrate (NaNO₃), urea (CH₄N₂O), and a mixture of all these sources. Highest values for Maximum Sustainable Yield (MSY), a measure of potential harvest rate based on population productivity, were observed in the urea treatment, but cell size was smaller compared to other treatments. Sodium nitrate and the mixture of nitrogen sources also had high MSY values but larger cell sizes, making them the treatments with highest total biovolume. The highest percentages of lipid by weight, but also highest densities of invading organisms were observed in the mixed treatment. Our results suggest that tradeoffs between biovolume and lipid yields as well as culture success can ultimately decide what nitrogen sources to use.     

Comparison of pyrolysis and hydrothermal liquefaction of Chlamydomonas reinhardtii. Growth studies on the recovered hydrothermal aqueous phase

The production of bio-oil by pyrolysis with a high heating rate (500 K s⁻¹) and hydrothermal liquefaction (HTL) of Chlamydomonas reinhardtii was compared. HTL led to bio-oil yield decreasing from 67% mass fraction at 220 °C to 59% mass fraction at 310 °C whereas the bio-oil yield increased from 53% mass fraction at 400 °C to 60% mass fraction at 550 °C for pyrolysis. Energy ratios (energy produced in the form of bio-oil divided by the energy content of the initial microalgae) between 66% at 220 °C and 90% at 310 °C in HTL were obtained whereas it was in the range 73–83% at 400–550 °C for pyrolysis. The Higher Heating Value of the HTL bio-oil was increasing with the temperature while it was constant for pyrolysis. Microalgae cultivation in aqueous phase produced by HTL was also investigated and showed promising results.     

Enhancement of lipid productivity with a novel two-stage heterotrophic fed-batch culture of Chlorella protothecoides and a trial of CO2 recycling by coupling with autotrophic process

In this study, the influences of oxygen supply and nitrogen starvation on cell growth and lipid production by heterotrophic culture of Chlorella protothecoides were investigated in a 5-L stirred-tank bioreactor. The results demonstrated that both a low oxygen supply and nitrogen starvation are favorable for lipid accumulation, but limit cell growth and may therefore reduce the overall lipid production. Hence, a novel two-stage strategy for oxygen supply and nitrogen starvation was proposed in a batch culture, increasing the lipid content and yield on glucose by a factor 2.14 and 2.03 compared to the one-stage batch culture with high oxygen supply and sufficient nitrogen availability, respectively. Applying the two-stage process in a fed-batch culture further increased the lipid productivity to 175.2 mg/(L·h), giving a lipid yield of 195.6 mg/g. Furthermore, a dual-mode culture, coupling a heterotrophic process with autotrophic process, was implemented to allow for CO₂ recycling. This improved the lipid productivity and carbon utilization efficiency, further contributing towards cost-effective and environment-friendly algal lipid production.     

Hydrogen production by Chlamydomonas reinhardtii under light-driven and sulfur-deprived conditions: Using biomass grown in outdoor photobioreactors at the Yucatan Peninsula

Photosynthesis is the ultimate natural process that supports all the biofuels generation. Photosyntetic production of hydrogen by microalgae is very attractive from the renewability point of view. Moreover, it faces several challenges: since the process itself has a low yield, a large number of considerations should be studied to optimize the hydrogen production at the lowest cost. In this work, wild-type Chlamydomonas reinhartii was grown outdoors in the Yucatán peninsula. Three different diameters of tubular photobioreactors (PBRs), two autotrophic culture media, as well as two seasons of the year were analyzed. From these variables, it was determined that the best biomass yield was during the winter season and with the Sueoka culture medium. Statistical significance differences were not found for the diameters of the PBRs. During growth, the biomass was exposed to natural light–dark cycles and at the end of the exponential phase of growth it was harvested with superabsorbent polymers. This biomass was able to produce hydrogen under anaerobic conditions in Tris-Acetate-Phosphate culture medium in indoor PBRs exposed to continuous artificial illumination. Experiments with different initial biomass concentrations in the anaerobic PBRs showed direct relationship with the hydrogen production profile.     

One-step multi enzyme pretreatment and biohydrogen production from Chlorella sp. biomass

One-step multi-enzyme (OSME) pretreatment of Chlorella sp. biomass were statistically optimized. Enzyme dosages of 380 units/g-VS Termamyl SC, 229.45 units/g-VS Dextrozyme GA, and 34.13 filter paper units/g-VS Cellic CTec 2, and a hydrolysis time of 90 min were the optimal conditions yielding a RSY of 167.61 mg/g-VS, equivalent to 49.29% carbohydrate solubilization. An OSME pretreated biomass was used as substrate for biohydrogen production that was optimized under feed/inoculum ratio of 3.0 and initial pH of 6.5. A hydrogen yield of 43.16 mL-H₂/g-VS was obtained under optimal conditions, which was 39.63% greater than that observed with non-treated biomass. Addition of nutrients in the form of a basic anaerobic medium with or without a nitrogen source, tended to decrease the efficiency of hydrogen production from the hydrolyzed biomass. This work demonstrated OSME hydrolysis as an alternative to the sequential hydrolysis processes for efficient conversion of microalgae biomass to fermentable sugars and biohydrogen.     

Oxygen and nitrogen-doped metal-free microalgae carbon nanoparticles for efficient hydrogen production from sodium borohydride in methanol

Here, the oxygen(O) and nitrogen(N) doped metal-free carbon synthesis including potassium hydroxide (KOH) activation of Spirulina Platensis microalgae, followed by nitric acid (HNO₃) activation is reported for the first time. Oxygen and nitrogen-doped metal-free catalysts were investigated for efficient hydrogen (H₂) production from methanolysis of sodium borohydride (NaBH₄). Compared to the catalyst obtained with the KOH activation, the catalytic activity for O and N doped metal-free showed about a four-fold improvement. The catalysts were analysed by scanning electron microscope (SEM), X-ray photoelectron spectroscopy (XPS), X-ray diffractometer (XRD), nitrogen adsorption, elemental analysis and Fourier-transform infrared spectroscopy (FTIR). The effects of temperature, NaBH₄ amounts, catalyst loading and reusability experiments on the catalytic performance of obtained metal-free catalysts by H₂ release from NaBH₄ methanolysis were performed. This study can provide a new alternative strategy to produce specific metal-free carbon catalysts doped heteroatom for environmentally friendly conversion to produce H₂ efficiently.     

Sustainable biohydrogen production by Chlorella sp. microalgae: A review

The use of fossil fuels is causing a huge environmental impact due to the emission of air pollutants, greenhouse gases, and other ground and water contaminants; also, these fuels are depleting; the world is facing an energy crisis in the years to come if no preventive actions are done. Renewable energies are arising as promising technologies that will complement and even replace conventional fuels shifting the global energy matrix to a cleaner and eco-friendly future. Microalgal biohydrogen is one of those emerging technologies that is showing positive results. This work provides an overview of the key parameters to produce hydrogen from microalgae especially from the genus Chlorella. Current status of chemical and biological hydrogen producing technologies is presented, along with the main metabolic processes for this purpose in microalgae, their characteristic enzymes, several strategies to induce hydrogen production, the key operation parameters and finally providing some remarks about scaling-up and industrial-scale applications.     

Sulphur and nitrogen-doped metal-free microalgal carbon catalysts for very active dehydrogenation of sodium borohydride in methanol

Micro algae based on Spirulina platensis is successfully used for the synthesis of S and N-doped metal-free carbon materials. The procedure consists of three stages; (i) Activated carbon production by KOH activation in CO₂ atmosphere (S-AC), (ii) S atom doping to the obtained S-AC using sulphuric acid by hydrothermal activation (S-AC-S), (iii) N atom doping by hydrothermal activation to S-AC obtained using nitric acid (S-AC-S-N). The S and N doped metal-free catalysts are used for H₂ release in NaBH₄ methanolysis reaction (NaBH₄-MR) for the first time. The metal-free carbon catalysts are characterized by X-ray photoelectron spectroscopy (XPS), scanning electron microscope (SEM-EDS), X-ray diffractometer spectroscopy (XRD), Fourier-transform infrared spectroscopy (FTIR), nitrogen adsorption and elemental analysis (CHNS) methods. When the HGR values obtained for S-AC-S-N (26,000 mL min^?1 g^?1) and S-AC (2641 mL min^?1 g^?1) are compared, there is a 9.84-fold increase. Activation energy (Ea) value for S-AC-S-N was 10.59 kJ mol^?1.     

Chlorella vulgaris microalgae strain modified with zinc chloride as a new support material for hydrogen production from NaBH4 methanolysis using CuB,NiB, and FeB metal catalysts

This study aims to produce hydrogen (HG) from sodium borohydride (NaBH₄) methanolysis using CuB, NiB or FeB catalysts prepared with a different support material including C. vulgaris microalgae strain modified using zinc chloride (CMS-ZnCl₂). The NaBH₄ concentration, metal percentage in the supported-catalyst, the optimal ZnCl₂ percentage, and temperature effect on the NaBH₄ methanolysis were investigated. X-ray photoelectron spectroscopy (XPS), X-ray powder diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), Transmission electron microscopy (TEM), and Scanning electron microscopy (SEM) analysis were performed for the CMS-ZnCl₂-CuB characterization. Also, the low activation energy (Ea) of 22.71 kJ mol⁻¹ was found with the supported catalyst obtained. Under the same conditions, nearly 100% conversion was achieved in the reusability experiments repeated five times, but a gradual decrease in catalytic activity was observed after each use.     

Metal-free catalysts with phosphorus and oxygen doped on carbon-based on Chlorella Vulgaris microalgae for hydrogen generation via sodium borohydride methanolysis reaction

Metal-free catalysts (C–KOH–P) containing phosphorus (P) and oxygen (O) prepared by the modification with phosphoric acid (H₃PO₄) of activated carbon (C–KOH) obtained by activation of Chlorella Vulgaris microalgae with potassium hydroxide (KOH) were investigated for the hydrogen (H₂) generation reaction from methanolysis of sodium borohydride (NaBH₄). Elemental analysis, XRD, FTIR, ICP-MS, and nitrogen adsorption were used to analyze the characteristics of metal-free catalysts. The results showed that groups containing O and P were attached to the carbon sample. In the study, the hydrogen production rates (HGR) obtained with metal-free C–KOH and C–KOH–P catalysts were 3250 and 10,263 mL/min/g, respectively. These HGR values are better than most values obtained for many catalysts presented in the literature. Besides, relatively low activation energy (Ea) of 27.9 kJ/mol was obtained for this metal-free catalyst. The C–KOH–P metal-free catalyst showed ideal reusability with 100% conversion and a partial reduction in the H₂ production studies of NaBH₄ methanolysis after five consecutive uses.