REVIEW ARTICLE

Microalgae: A Promising Source of Biofuels

Tulsi Ram Dadsena¹, Janavy Sahu¹, Sakshi Agrawal¹, Arvind Agrawal²*

¹School of Studies in Life Sciences, Pt. Ravishankar Shukla University, Raipur (CG) 492 010, India

²UGC-MMTTC, Pt. Ravishankar Shukla University, Raipur (CG) 492 010, India

Authors Email- tulsiramdadsena10@outlook.com; janavysahu@gmail.com; sakshi01agrawal01@gmail.com; dr.arvind02@gmail.com

*Corresponding Author Email- dr.arvind02@gmail.com

1.       Introduction

It is now widely accepted that using fossil fuels is not environmentally viable due to their enormous consumption and owing to the rise in CO₂ and other pollutants, such as heavy metals, in the atmosphere (Chisti 2007). The increased usage of fossil fuels during the Industrial Revolution has led to a notable rise in greenhouse gas (GHG) emissions, which has resulted in global warming and air pollution. Major cities like New Delhi, Beijing, Lima, and Karachi are seeing a rise in air pollution. The atmosphere is subjected to millions of tons of fluorinated chemicals, mercury, freon, and other harmful pollutants, as well as 150 million tons of SO2, 53 million tons of nitrogen oxides, and 20 billion tons of CO2 annually (Liu and Li 2023). 90% of people living in cities are exposed to air pollution, with PM 2.5 levels over the WHO's recommended level of 10 μg/m³. The International Agency for Cancer Research categorized external air contamination as a human carcinogen in 2013. Degraded atmospheric conditions and air pollution are linked to heart diseases, stroke, acute and chronic respiratory diseases, and lung cancer. In 2016, 7 million people lost their breaths from household and ambient air pollution. Indoor air pollution results in the deaths of around 2.5 million individuals annually across the globe. Fossil fuel resources accounts for 75% of emissions of heat-trapping gases on a global scale and contributes heavily to local air pollution, causing 5 million premature deaths each year. Following the Industrial Revolution, the majority of industries were primarily driven by steam and diesel engines, with fossil fuels serving as the primary energy source. Between 2000 and 2019, global CO2 emissions rose at the fastest pace recorded in human history, contributing to 30.7% of total global emissions. Observational data indicates that the global mean surface temperature (GMST) has been increasing, and this pattern has been distinctly evident since 1950. In 1880, the GMST was −0.16˚C; by 2021, it had risen to 0.84˚C, an increase of 1.0˚C. The average surface temperature in the Northern Hemisphere rose by 1.42˚C from −0.28˚C in 1880 to 1.14˚C in 2021. The average surface temperature in the Southern Hemisphere rose by 0.59˚C from −0.04˚C in 1880 to 0.55˚C in 2021. The GMST of 1.02˚C in 2020 was the highest temperature ever documented (Liu and Li 2023).

Concerns about the scarcity of fossil fuels, the rising cost of crude oil, energy security, and the increase of global warming have sparked a global interest in biofuels and other renewable energy sources. Carbon-neutral and renewable transport fuels are vital for sustaining both the economy and the environment (Chisti 2007). Utilizing unrefined vegetable oils as fuel might cause multiple engine issues, including carbon deposits on the engine's head and piston, choking of the injectors, and excessive engine wear. Many researchers have suggested using transesterified oils, also known as biodiesel, to solve these issues as they can significantly reduce the viscosity of the oil. Long-chain triglycerides from oils undergo a process known as transesterification, producing fatty acid methyl esters, referred to as FAME, which is the term for biodiesel. The two most widely utilized forms of biofuel are biodiesel and bioethanol, which are mostly made from lignocelluloses, seeds, and vegetable or microbial oils. Diesel may be replaced with biodiesel, and petrol can be replaced with bioethanol. Green hydrogen has also been used as the alternative of fossil fuels; Dark fermentation has been shown as a most efficient technology for the production of the green hydrogen (Vaswani et al. 2025). Additionally, water hyacinth has also been successfully tested for the biohydrogen by Tandon et al. (2019a) due to its high energy content.

2.       Generations of biofuels

Biofuels are divided into four forms as: first, second, third, and fourth-generation biofuels (Malode et al. 2022).

1.       First-generation biofuels

First-generation biofuels are derived from edible resources including sugar, starch, vegetable oil (soybean, mustard, coconut, palm, corn, rice, rapeseed, olive, and so forth), and animal fats using traditional technology. The main benefits of the raw material from the first-generation are its easily accessible yields and very straightforward transformation process. Biodiesel, sometimes known as fatty acid methyl esters chemically, is a fuel that is widely utilized in European nations and is comparable to mineral diesel.

The limitations of the first-generation biofuels include inability to adapt to different environmental conditions, high cost, and limited development area prevent biodiesel from being produced from edible raw materials. In 2006, China and India accounted for around 11 percent of the global ethanol production, using a combination of wood, maize, and other starches as feedstocks. Furthermore, when ethanol increases in fuel balance, the amount of fossil fuel byproducts produced by the photosynthetic carbon uptake by corn plants decreases.

The most prevalent worry about first-generation biofuels today is that, as production capacities rise, they will compete with agriculture for arable land needed to produce food. There may be significant food shortages as a result of the growing demand on arable land now utilized for food production, particularly within low- and middle-income nations where more than 800 million individuals already face hunger and malnutrition. Furthermore, heavy irrigation, fertilizer, and pesticide use, along with intensive land use, can lead to serious environmental issues.

Fig. 1: Technique employed in the synthesis of first-generation biofuels (Malode et al. 2022)

2.       Second-generation biofuels

Another name for the second-generation biofuels is advanced biofuels. With the introduction of second-generation biofuels. Fuels are meant to be produced from the woody portion of plants known as lignocellulosic biomass, which does not interfere with food production. The non-edible parts of corn or sugarcane, as well as agricultural residues, forest harvesting residues, or wood processing waste like leaves, straw, wood chips, or rice straws using Saccharomyces cerevisiae (Verma et al., 2020) are some of the sources. These fuels are created via biochemical conversion mechanisms or thermochemical reactions. The common thermochemical processes for converting biomass into synoil and syngas are pyrolysis, gasification, and liquefaction.

Second-generation biofuels, however, cannot yet be generated profitably on a large scale due to the requirement of expensive technologies involving pre-treatment with certain enzymes with the goal of transforming the woody biomass into fermentable sugars (Dragone et al., 2010). The necessity for land, water, and other resources to produce feedstock for biofuels results in a number of negative outcomes. The alteration in land use could lead to increased release of greenhouse gases and greater pressure on water supplies, pollution of the air and water, and increased food prices. Biofuels can produce even greater climate-impacting emissions compared to certain fossil fuels, Subject to variations in feedstock and the procedure used. The lignocellulosic waste has been also used by Tandon et al. (2019b) and pH and temperature has been optimized for the maximum yield.

Fig. 2: Process of production of second-generation biofuels (Malode et al. 2022)

3.       Third-generation biofuels

Third-generation biodiesel is the term for the biodiesel made from leftover oils and microalgae. The main sources of third-generation biodiesel include waste cooking oil, fat, fish oil, microalgal oil and its derivatives. Microalgal species can accumulate high lipid content, making them a possible future hotspot for third-generation biodiesel. Cyanobacteria, and microalgae are examples of microorganisms that produce third-generation biofuels. In a stressful environment, oleaginous bacteria and microalgae start to store lipids in their bodies serving as an energy provider. Catalysis and transesterification of extracted lipid and short chain alcohol may assist in manufacturing of biodiesel. Third-generation biodiesel has several important benefits, including less of an influence on nurseries, increased development order and efficiency, a higher oil rate, and less of an impact on food delivery. The abundance of potential benefits associated with third-generation biodiesel overwhelms the drawbacks of previous generation feedstocks, including their influence on the natural hierarchy, accessibility, ecological boundary flexibility, and financial feasibility.

Microalgae are regarded as the ideal option for producing biofuels and synthetics due to their high photosynthetic transformation proficiency, quick development, less land requirements, and resilience to harsh growth situations. Carbohydrates, and hydrocarbons are the carbon sources that oleaginous microorganisms (such as yeast, fungus, and microalgae) can use to make microbial oil, sometimes referred to as single cell oil. Microbial oil is mostly composed of triglycerides composed of PUFA. On the other hand, highly unsaturated fatty acids, such as C16 and C18, share chemical similarities with vegetable oils, such as soybean, palm, and rapeseed oils. These suggest that biodiesel made from microbial oil should have a generally acceptable grade. According to microbiology, the oleaginous microorganisms that produce lipid are classified into three distinct categories family groups: bacteria, fungi (such as Molds and yeast), and microalgae. Of these, bacteria are less competent of creating lipid since they can only produce certain lipid and PUFA. That is why, microalgae and fungi are acknowledged to be the key lipid producers.

Harnessing microalgae for the purpose of biofuels production offers the several benefits when compared to higher plants (Dragone et al., 2010):

(1) Besides their rapid growth, Microalgae produce and store a considerable quantity of neutral lipids (20–50% dry mass of biomass);

(2) Microalgae cultivation is feasible throughout the entire year under controlled or suitable environmental conditions, the oil yield of microalgae cultures per unit area may surpass that of the best oilseed crops;

(3) They require less water than terrestrial crops, freshwater resources are not as burdened;

(4)  There is no requirement for pesticides or herbicide in the microalgae cultivation;

(5) With a significant lipid content, microalgae are excellent carbon concentrators (Singh et al., 2024). Microalgae reduce significant greenhouse gases by sequestering CO₂ from flue gases released by coal-powered energy plants and other sources (1-kilogram dry biomass of algae utilises approximately 1.83 kilograms of CO₂);

(6) Removal of NH₄⁺, NO₃^-, and PO₄^3- from a range of wastewater sources as part of effluent bioremediation.

(7) Since microalgae may be grown in saline, brackish, or coastal waters on non-arable soil and can withstand stressful conditions and require less nutrients, they do not compete with conventional agriculture for resources;

4.       Fourth-generation biofuels

In fourth-generation biofuels, biomass is produced from crops or microalgae that have been genetically modified to absorb substantial quantities of carbon. After that, the crops are turned into fuel. Because the carbon involved in their production is removed from the environment, several of these fuels are regarded as carbon negative. However, for a variety of reasons, including a paucity of genomic data, the complexity of transgenesis, and challenges in achieving a competence equilibrium between metabolic and energy storage pathways, genetic alteration is not currently feasible for all algal species. The simplest approach to lower costs, nutrient usage, and water footprint is to increase production and lipid accumulation. The productivity and lipid content of microalgae can be increased through the widespread use of genome editing techniques.

Currently two different approaches have been used for the fourth-generation biofuels:

1.       Metabolic pathway engineering in microorganisms, including cyanobacteria and microalgae, involves optimizing growth rates, utilizing diverse carbon sources, and channelling metabolic flux toward enhanced biofuel synthesis and elevated production efficiency.

2.       Engineering of model organisms with biosynthetic pathways originally present in natural producers, leveraging their genetic accessibility.

A study conducted in Europe concluded that most customers would accept genetically modified algae for synthesizing biofuels if the systems' safety could be ensured (Cavelius et al., 2023).

Fig. 3: Process for production of Third-generation Biofuels (Dragone et al. 2010)

Table 1: Some of the manipulation techniques used in microalgae utilized for the synthesis of fourth-generation biofuels (Moravvej et al., 2019)

Table 2: Comparative Analysis of Various Biodiesel Feedstocks (Chisti 2007)

^a To supply half of the transportation fuel required in the United States.

^b 70% oil (by weight) in biomass.

^c 30% oil (by weight) in biomass.

To supply merely half of the US's transportation fuel needs, 24% of all cropland must be planted with oil palm, known for its high oil yield. It is obvious that oil crops will not be able to substantially replace liquid fuels derived from petroleum in the near future. If biodiesel is made from microalgae, this scenario is drastically altered. It would take nearly 1 to 3% of all U.S. cropland to produce enough algal biomass To fulfil half of the country's transportation fuel requirements (Chisti 2007).

The majority of the ongoing study is concentrated on a few fast-growing microalgal species that have been shown to accumulate significant amounts of lipids, albeit under certain circumstances. The common green algae species are Scenedesmus, Chlamydomonas reinhardtii, Dunaliella salina, Botryococcus braunii, (which grows slowly but can collect huge amounts of lipids), and several Chlorella species. Although many microalgae strains have a high lipid content, it is feasible to boost that concentration by optimizing parameters that affect growth, such as temperature, salinity, light intensity, nitrogen level control, CO₂ concentration, and harvesting technique.

3. Factors affecting quality of microalgal oils

The culture parameters of holophytic microalgae, including light intensity, temperature, pH, carbon dioxide supply, culture medium composition, growth cycle, heavy metal concentration etc., have a direct impact on microalgal oil quality (Ma et al., 2018)^.

1.       Effect of light intensity:

Three variables of light important to enable optimal microalgal growth are: 1. Intensity 2. Spectral quality 3. Photoperiod.

According to studies, green algae grow more readily under blue and red light because these wavelengths are favoured by the pigments chlorophyll a and b, which are important light-harvesting pigments (Singh and Singh, 2015).

Triglycerides and other non-polar lipids are commonly used to make biodiesel. The synthesis of non-polar lipids is favoured by high light intensity, whereas the synthesis of polar lipids is favoured by low light intensity. Strong light preferentially increases the triacylglycerol (TAG) content (Maltsev et al., 2021). The light conditions in natural environments are highly varied and depend on a range of factors, including the geographical latitude of the region, temporal (daily and seasonal) variations in the solstice altitude, light scattering by the atmosphere, water column, etc. Accordingly, microalgae are adapted to various lighting conditions. Some species are found in highly shaded habitats, for example, in the soil under the dense canopy of higher plants, above-water and underwater caves, in deep water layers, and the other species are adapted to grow in direct sunlight, for example, in polar, tropical deserts, on salt marshes and various other open surfaces, where the light intensity can reach 2000 µmol m⁻²s⁻¹. At low light levels, the rate of photosynthesis increases almost linearly with increasing light intensity until the region of saturating light is reached. A continued rise in the light intensity may lead to the photo inhibition or ROS-mediated cellular injury to the photosynthetic apparatus. Photon absorption is facilitated by photopigments. A high-energy state is achieved by the excited pigment molecules upon absorption of a light quantum. Photochemical changes take place because they release energy when they return to their original condition. Each microalgal photosystem contains a different set of light-harvesting pigments that provide a unique absorption spectrum.

Table 3: Optimum light intensity values for the maximum growth rate of algae from different taxonomic groups (Maltsev et al., 2021).

2.       Effect of salinity:

Change in salinity causes osmotic stress, ion stress and changes of cellular ion concentration. The algal cells can withstand a lower NaCl concentration, allowing them to proliferate normally. However, when the NaCl concentration rises, chlorophyll breakdown occurs, which results in cell death. Hence, the microalga must have an optimum NaCl concentration to grow properly; growth rate decreases at higher or lower concentrations (Rai et al., 2015). Marine microalgae have extreme tolerance to changes in salinity. Dunaliella sp., a marine microalga, grows with a salinity range of 0.05–5.5 M NaCl (Mohanta et al., 2023). Researches on photosynthesis shows that the Hill reaction, which produces adenosine triphosphate and flavin mononucleotide, requires chloride ions. It's interesting to note that autotrophically growing Chlorella sp. need higher chloride ions, as demonstrated by Eyster (1958). The study conducted by Ben‐Amotz et al. (1985) revealed that Isochrysis sp. cells cultured at 29 ppt had a differing profile of fatty acids from those grown at 58 ppt. The cells had lower levels of polyunsaturated C18 and C22 fatty acids, while some C16 and C18 fatty acids were higher.

3.       Effect of temperature and pH:

The range of 16 to 27°C is ideal for the majority of microalgae growth; for example, microalgae growth tends to slow down at conditions where the temperature is less than 16°C and may be significantly impeded Under temperatures exceeding 35°C. The degree of unsaturation of the fatty acids generated generally has an inverse relationship with the culture temperature. For instance, at 25°C, Chlorella vulgaris had a 14.7% lipid content, while at 30°C, only roughly 5.9% was found (Ma et al., 2018). The cell can perform photosynthesis at ideal growth temperatures without altering any of its natural biochemical or physiological processes. Maximum growth rates for mesophilic species are specified to be between 20 and 25°C; however, for thermophilic strains (Chaetoceros, Anacystis nidulans), these temperatures can rise to 40°C, and for psychrophilic strains (Asterionella formosa), they can drop to 17°C.

The literature suggests that when studying the same species, the ideal growth temperatures can vary. The ideal temperature cursor may move in either direction depending on the environment around it. When the NaCl content was increased from 0.125 to 1.5 M, optimal growth temperature of Dunaliella tertiolecta increased by 6°C (Ras et al., 2013). All metabolic processes are influenced by temperature, and the ideal temperature for a specific strain of algae will significantly impact the culture's potential productivity. The photosynthesis rate, respiration and growth of microalgae all decline due to imbalances between adenosine triphosphate (ATP) production and energy demand, inactivation or denaturation of necessary proteins for photosynthesis or stress on photosystem II activity when optimal temperatures are exceeded. The fatty acid profile is also affected by temperature. Total unsaturated fatty acids have been shown to decrease at high temperatures (Chaisutyakorn et al., 2018).

The culture pH is another important factor which influences the enzymatic activity and synthesis of microalgae oils. In general, neutral or slightly acid conditions favor the microalgal growth. The pH value of the actively growing culture in the culture medium has to be maintained by simple aeration or supplementation with extra CO₂ since the pH increases with the growth of microalgae as a result of the continuous consumption of CO₂.

4.       Effect of carbon dioxide:

The Carbon dioxide concentration, an inorganic carbon source, controls the development and lipid accumulation of microalgae. Low concentrations of CO₂ can stimulate the microalgal growth while inhibiting the synthesis of fatty acids; conversely, high concentrations of CO₂ stimulate the synthesis of fatty acids while influencing the desaturation and extension of the carbon chain. Tsuzuki et al. (1990) reported that Chlorella vulgaris fatty acid composition changed in response to the CO₂ concentration during growth. When cells were grown in conditions with high CO₂, less desaturation was seen. Conversely, the CO₂ concentration had no measurable influence on the fatty acid chain length. This study highlights that, in addition to photosynthetic properties, membrane lipid composition is influenced by ambient CO₂ concentration. In different eukaryotic microalgae, de novo carbonic anhydrase synthesis is induced under low CO₂ concentrations, and a pyrenoid with a starch sheath develops. Compared to water, carbon dioxide is much more soluble in fats and adsorbs quickly. Thus, one of the reactions that arises in response to the decrease in CO₂ concentration is the change in the degree of unsaturation of the fatty acyl chain in the membranes.

5.       Effect of culture medium composition:

The inorganic components that make up an algal cell must be supplied by the growing medium. Nitrogen (N) and phosphorus (P) are examples of essential elements. The microalgal biomass's approximate molecular formula, CO_0.48H_1.83N_0.11P_0.01, (Chisti, 2007) can be used to calculate the minimum nutritional requirements. Most nitrogen is given as nitrate (NO₃^-), although urea and ammonia (NH₄⁺) are also frequently utilized. The best nitrogen source is urea because it produces larger yields at the same nitrogen content and results in less pH variations in the medium during algal development. Conversely, given that the added phosphates bind with metal ions, some of the added P is not bio-available and must be supplied in large excess (Dragone et al. 2010).

Depending on which nutrient is limited and to what extent, algae can exhibit significant variation in their biochemical composition when under conditions of nutrient limitation. Under ideal temperature and pH levels, the growth rate of algae is generally proportional to the uptake rate of the most limiting nutrient, and this relationship is typically represented by the Michaelis-Menten equation (Baiee and Salman, 2016). The lipid content and composition of autotrophic microalgae are significantly influenced by nitrogen and phosphorus in their culture medium. Of these, the effect of nitrogen is more evident on the fatty acid metabolism of autotrophic microalgae, as evidenced by the fact that nitrogen-limiting conditions can increase total lipid content. Most notably, when microalgae are exposed to alternating conditions of sufficient and limited nitrogen, the composition of their lipid changes from free fatty acids to triglycerides. Microalgal cells are supported by starch synthesis when the culture medium has a high nitrogen content. However, phosphorus in the culture media also plays a role in the production of fatty acids. Research has demonstrated that Scenedesmus obliquus can reach up to 30% of its total lipid content when it is subjected to phosphorus limitation, while it can reach up to 53% of its total lipid content when it is subjected to both phosphorus and nitrogen limitation simultaneously.

6.       Effect of growth cycle

The fatty acid composition and lipid content of microalgae can change as they grow through different phases. During the stationary phase, microalgae can reach their maximum lipid content. TAG content decreases in most microalgae during the logarithmic phase of growth and increases during the stationary phase, suggesting that TAGs serve as storage lipids (Khozin-Goldberg et al., 2005; Volkman et al., 1989). The growth phase of microalgae could be a useful variable for the content enrichment of essential fatty acid (EFA) rich lipids, according to a study conducted by Li et al. (2014) on six important diet microalgae (Chlorella sp., Nannochloropsis oculata, Nannochloropsis sp., Isochrysis galbana, Phaeodactylum tricornutum Bohlin and Chaetoceros calcitrans). For Nannochloropsis sp. and N. oculata, a higher content of eicosapentaenoic acid (EPA-rich) diacylglyceryl-trimethylhomoserine (DGTS) and a higher content of docosahexaenoic acid (DHA-rich) lipids could be obtained at the end of stationary phase, respectively. Nonetheless, EPA/AA-rich lipids were primarily found in TAG and monogalactosyl diacylglycerol (MGDG), respectively, in both P. tricornutum and C. calcitrans. Furthermore, for P. tricornutum, a higher concentration of EPA-rich TAG might be obtained at the end of the stationary phase. At the conclusion of the stationary phase, C. calcitrans may have a higher content of EPA-rich MGDG.

7.       Effect of Heavy metals:

Microalgae exhibit a high degree of sensitivity to environmental fluctuations. Even trace concentrations of various organic and inorganic pollutants, including heavy metals, can induce notable alterations in their overall metabolic activity. Owing to this sensitivity, microalgae are frequently employed as biological indicators for assessing the potential toxicity of heavy metals (Chouteau et al. 2004; Durrieu et al. 2011; Torres et al. 2000). Resistance to heavy metals in microalgae may stem from their capacity to limit metal uptake. This can be facilitated through the adsorption of toxic metal ions onto cell-associated substances or structural components of the cell wall, as well as through the secretion of metal-chelating organic compounds into the surrounding environment.

4. Production of biodiesel from microalgal oil:

Microalgal lipids (TAGs) can be directly converted into FAMEs using the process of transesterification (Griffiths et al., 2010).

   Fig. 4: Process of transesterification

Transesterification is a chemical process involving the exchange of organic alkyl groups between an ester and an alcohol (R1, R2, R3) of microalgal or vegetable/plant oil with the methyl group of methyl alcohol. Such reactions are typically facilitated by either acid or base catalysts.

Initiatives for biodiesel were launched in India in 2001 using second-generation feedstocks, such as inedible seeds like Jatropha. The National Mission on Biodiesel was established in 2003 to promote the manufacture of biodiesel by combining jatropha seeds with fuel. With a fixed price of Rs. 25/L in 2003 that rose to Rs. 26.50/L by 2008, the goal was to achieve 20% blending by 2011–2012.

5. Four types of microalgae cultivation based on mode of nutrition can be adopted:

1.       Photoautotrophic:

This pathway uses light and CO₂ to produce energy and cellular carbon i.e. photosynthesis. The requirement of carbon is supplied by inorganic carbon sources like CO₂, Na₂CO₃, NaHCO₃ etc.

2.       Heterotrophic:

In this pathway energy and carbon is produced from the organic carbon source like glucose and acetate and transformed into carbon intermediate in the main metabolic pathways. Several algal strains e.g. Chlorella protothecoides, Galdieria sulphuraria and Neochloris oleoabundans Have been investigated under heterotrophic conditions for enhanced biomass and fatty acid production. (Morales-Sánchez et al., 2015).

3.       Mixotrophic:

Under mixotrophic cultivation, photo-autotrophy and heterotrophy take place simultaneously because the microalgae use both organic carbon sources and inorganic CO₂ in the presence of light. While organic compounds are assimilated through aerobic respiration, which is influenced by the availability of organic carbon, CO₂ is fixed through photosynthesis, which is dependent on illumination. Mixotrophic cultivation has higher growth rates than either heterotrophic and photo-autotrophic regimes by shortening growth cycles, prolonged exponential growth phase and producing higher biomass (Lowrey et al., 2015).

4.       Photoheterotrophic:

Energy derived by photosynthesis is used for the effective metabolism of organic carbon source. the primary carbon source being organic carbon; decreased reliance on photosynthesis; and enhanced organic carbon metabolism through the use of light-driven ATP (Cheng et al.,2022).

Fig. 5: Types of mode of nutrition for the microalgal growth (Perez-Garcia & Bashan, 2015)