Dolly Wattal Dhar

Microalgae as second generation biofuel. A review

Microalgae are autotrophic microorganisms having extremely high photosynthetic efficiency and are valued as rich source of lipids, hydrocarbons, and other complex oils for biodiesel besides being an invaluable source of bioethanol, biomethane, and biohydrogen. Biodiesel produced from oilseed crops such as jatropha and soy have lower yields per unit land area and threaten food security. Indeed, microalgae have higher oil yields amounting to about 40 times more oil per unit area of land in comparison to terrestrial oilseed crops such as soy and canola. Further, microalgae production does not require arable land for cultivation. Biofuel is regarded as a proven clean energy source and several entrepreneurs are attempting to commercialize this renewable source. Technology for producing and using biofuel has been known for several years and is frequently modified and upgraded. In view of this, a review is presented on microalgae as second generation biofuel. Microalgal farming for biomass production is the biggest challenge and opportunity for the biofuel industry. These are considered to be more efficient in converting solar energy into chemical energy and are amongst the most efficient photosynthetic plants on earth. Microalgae have simple cellular structure, a lipid-rich composition, and a rapid rate of reproduction. Many microalgal strains can be grown in saltwater and other harsh conditions. Some autotrophic microalgae can also be converted to heterotrophic ones to accumulate high quality oils using organic carbon. However, there are several technical challenges that need to be addressed to make microalgal biofuel profitable. The efficiency of microalgal biomass production is highly influenced by environmental conditions, e.g., light of proper intensity and wavelength, temperature, CO2 concentration, nutrient composition, salinities and mixing conditions, and by the choice of cultivation systems: open versus closed pond systems, photobioreactors. Currently, microalgae for commercial purpose are grown mostly in open circular/elongated “raceway” ponds which generally have low yields and high production costs. However, a hybrid system combining closed photobioreactor and open pond is a better option. The biggest hurdle in commercialization of microalgal biofuel is the high cost and energy requirement for the microalgal biomass production, particularly agitation, harvesting, and drying of biomass. In order to conserve energy and reduce costs, algae are often harvested in a two-step process involving flocculation followed by centrifugation, filtration, or micro-straining to get a solid concentration. However, the major bottlenecks in algal biodiesel production within the cell can be identified and handled by adopting a system approach involving transcriptomics, proteomics, and metabolomics. Research and developments in the field of new materials and advanced designs for cultivation in closed bioreactors, use of waste water for biomass production, screening of efficient strains, high-value coproduct strategy, and cutting-edge metabolic engineering are thought to provide the biggest opportunities to substantially improve the cost effectiveness of such production systems.

Biochemical modulation of growth, lipid quality and productivity in mixotrophic cultures of Chlorella sorokiniana

The genus Chlorella is a widely employed microalga for biodiesel, as it can be grown using photo/mixo/heterotrophic mode of cultivation. The present investigation was undertaken with the hypothesis that addition of different substrates (amino acids, carbon sources, vitamins) along with reducing agents may aid in diverting Acetyl CoA to malonyl CoA or fatty acid biosynthesis, under mixotrophic conditions in Chlorella sorokiniana. Preliminary investigations undertaken with two reducing agents individually (sodium thiosulphate and methyl viologen) along with selected substrates revealed the promise of sodium thiosulphate (1%) in enhancing lipid accumulation significantly. Further, the role of inclusion of twelve substrates and sodium thiosulphate revealed that supplementation with tryptophan (0.1%) recorded 57.28% enhancement in lipid productivity on 4th day. Highest values of lipid productivity of 33% were recorded on 8th day in 0.1% glucose supplemented medium containing sodium thiosulphate. Fatty Acid Methyl Ester (FAME) profiles generated revealed significant reduction in the content of Poly unsaturated fatty acids (PUFA) and enhanced Mono unsaturated fatty acids (MUFA) (especially oleic acid) in the treatments involving tryptophan, Vitamin B12, sodium pyruvate and glucose. This study reveals the promise of using sodium thiosulphate along with selected substrate for enriching the quality and quantity of lipids, which can be valuable for exploiting algae as a source of biodiesel.

Bioprospecting and indexing the microalgal diversity of different ecological habitats of India

Our study reports the collection, biodiversity analyses, isolation and identification of microalgae from different habitats of India. Cyanophyceae and Chlorophyceae were the most dominant algal groups recorded, with the highest number being recorded for non-heterocystous cyanobacteria (48), followed by 44 unicellular forms. Sagar Island, Sunderbans recorded the greatest number of algae, and unicellular/colonial green algae were present in all the samples. Shannon’s Diversity Index was highest in Koikhali, Sunderbans, followed by Rushikulya River, Odisha. Selective enrichment, purification through serial dilution followed by plating and regular observations led to the isolation of sixteen strains. Identification was done by using microscopic observations, supported with standard monographs and classified as belonging to seven genera (Chlorella, Chlorococcum, Kirchneria, Scenedesmus, Chlamydomonas, Tetracystis and Ulothrix). 18S rDNA sequencing was undertaken for four strains. The set of sixteen strains were screened under standard cultural conditions for their growth kinetics and Chlorella sorokiniana MIC-G5, followed by Chlorella sp. MIC-G4 exhibited the highest growth rates. The strain Chlorococcum sp. MIC-G2 recorded highest chlorophyll, while MIC-G3 ranked highest for carbohydrates. The study aided in identifying the dominant microalgae in the diverse habitats and characterizing their growth rate and carbohydrate content, providing a valuable germplasm for further utilization in agriculture and industry.

Comparative Performance of Three Carrier Based Blue Green Algal Biofertilizers for Sustainable Rice Cultivation

ABSTRACT Nitrogen fixing cyanobacteria or blue green algae are ecologically significant inputs in rice cultivation in the tropics. Field experiments were conducted to compare the efficiency of two newly developed carrier based blue green algal (BGA) biofertilizers (wheat straw and multani mitti), with the traditional soil based BGA biofertilizer, on the grain yield of rice for a period of three years. Treatments included five levels of nitrogenous fertilizer urea and their interaction with the three types of BGA biofertilizers, on the grain yield of rice variety ‘PNR 381’. Highest grain yields were obtained with the application of multani mitti based biofertilizer along with 90 kg N/ha, although maximum percent increase in yield over control (37.97%), when applied along with 60 kg N/ha. The straw based and soil based biofertilizer treatments showed highest yields when supplemented with 90 and 120 kg N/ha, respectively. This investigation clearly emphasizes the need for supplementing chemical fertilizers with the newly developed BGA biofertilizers in rice cultivation for maximizing crop productivity, reducing inputs of chemical fertilizers and sustaining soil fertility.

Cell disruption methods for improving lipid extraction efficiency in unicellular microalgae.

Identification of cost‐effective cell disruption methods to facilitate lipid extraction from microalgae represents a crucial step in identifying promising biofuel‐producing species. Various cell disruption methods including autoclaving, microwave, osmotic shock, and pasteurization were tested in the microalgae Chlorococcum sp. MCC30, Botryococcus sp. MCC31, Botryococcus sp. MCC32, and Chlorella sorokiniana MIC‐G5. Lipid content (on dry weight basis) from the four cultures on day 7 ranged from 11.15 to 48.33%, and on day 14 from 11.42 to 44.26%. Among the methods tested, enhanced lipid extraction was achieved through osmotic shock (15% NaCl) for Botryococcus sp. MCC32, microwave (6 min) for Botryococcus sp. MCC31, osmotic shock (5% NaCl) for Chlorella sorokiniana MIC‐G5 and microwave (2 min) for Chlorococcum sp. MCC30. The highest palmitate (16:0) contents (25.64% and 34.20%) were recorded with osmotic shock (15% NaCl) treatment for Botryococcus sp. MCC32 and microwave (6 min) for Botryococcus sp. MCC31, respectively. Two strains, along with their respective cell disruption methods, were identified as promising oil blends or nutraceuticals due to their high unsaturated fatty acid (UFA) content: Botryococcus sp. MCC31 (37.6% oleic acid content; 39.37% UFA) after autoclaving and Botryococcus sp. MCC32 after osmotic shock of 15% NaCl treatment (19.95% oleic acid content; 38.17% UFA).

Cyanobacterial Reclamation of Salt-Affected Soil

Salinity has been an important historical factor which has influenced the life span of agricultural systems. Around 10% of the total cropped land surface is covered with different types of salt-affected soils and the Asian continent accounts for the largest area affected by the salinity of various intensities. Cyanobacteria are capable of not only surviving, but thriving in conditions which are considered to be inhabitable, tolerating desiccation, high temperature, extreme pH and high salinity, illustrating their capacity to acclimatise to extreme environments. Until recently, the responses of cyanobacteria to salinity stresses were poorly documented as compared to heterotrophic bacteria and phototrophic eukaryotic algae. Cyanobacteria can be used to reclaim alkaline soils and fertility can be improved for subsequent cultivation of cereal crops, sugarcane and horticultural crops. Therefore we present here a review on cyanobacterial reclamation of salt-affected soil. Substantial progress has been made towards better understanding of the physiological mechanisms responsible for salinity tolerance and osmotic adjustment in cyanobacteria. Many researchers throughout the world have worked on probable mechanisms of salt tolerance studies in cyanobacteria. These organisms evolved about 3,000 million years ago and are considered to be the primary colonisers of the inhospitable ecosystems. The physiological aspects for the adaptation of cyanobacteria to high salinities include (a) synthesis and accumulation of osmoprotective compounds, (b) maintenance of low internal concentrations of inorganic ions and (c) expression of a set of salt-stress proteins. Exposure of cyanobacterial cells to different abiotic stresses resulted in rapid expression of several stress-regulated proteins and modifications in protein synthesis programme. The synthesis of organic solutes like disaccharides (sucrose, trehalose and glucosyl glycerol), quaternary amines (glycine betaine) and free amino acids (glutamine) are well-documented. The protection against alkaline environment is provided by the synthesis of specific fatty acids, sucrose- and osmotic-stress-induced proteins. In cyanobacteria, accumulation of internal osmoticum in the form of inorganic ions and prevention of intracellular Na+ accumulation by the curtailment of Na influx and by efficient active efflux mechanisms or metabolic adjustments have been investigated in depth. The Na+ extrusion in cyanobacteria is driven by a Na+/H+ antiporter, which is energised by enhanced activity of cytochrome oxidase. The inhibition of sodium ion influx appears to be a major mechanism for the survival of cyanobacteria against salt stress and synthesis of salt-stress proteins have been found in cyanobacteria. These organisms have been recognised as an important agent in the stabilisation of soil surfaces primarily through the production of extracellular polysaccharides which are prominent agents in the process of aggregate formation and increase in soil fertility. Cyanobacterial application results in the enrichment of soil with fixed nitrogen, soil structure improvement and declining trend of pH, electrical conductivity (EC) and Na+. The extracellular polysaccharides excreted by cyanobacteria have been reported to be responsible for binding of soil particles, thus, leading to the formation of a tough and entangled superficial structure that improves the stability of soil surface and protects it from erosion. The potential impact of these organisms on agriculture through their use as soil conditioners, plant growth regulators and soil health ameliorators has been well-recognised. Besides bringing about an improvement in the yield of rice, cyanobacteria produce direct and indirect beneficial changes in the physical, chemical and biological properties of soil and soil–water interface in the rice fields, which are of agronomic importance. Certain cyanobacteria have been found not only to grow in saline ecosystems but also improve the physico-chemical properties of the soil by enriching them with carbon, nitrogen and available phosphorus. Flushing of field may not be effective for the reclamation of saline soils and the addition of cyanobacterium inoculum along with the addition of gypsum is required before irrigation to ameliorate saline soils. Nitrogen-fixing cyanobacteria can be used as biological input to improve soil texture, conserve moisture, scavenge the toxic sodium cation from the soil complex and improve the properties of soils. Virtually negligible information exists on the genetics of cyanobacterial halotolerance. The presence of combined nitrogen which effectively curtails sodium accumulation and supports extra nitrogen demand for osmoregulation during slat stress confers considerable salt tolerance on cyanobacteria.

Biodiversity and biological degradation of soil

Soils contain enormous numbers of diverse living organisms assembled in complex and varied communities. Microscopic examination of a soil sample reveals the presence of billions of organisms like nematodes, protozoa, fungi, algae, actinomycetes, bacteria and cyanobacteria. These diverse organisms interact in the ecosystem, forming a complex web of biological activity. Environmental factors, such as temperature, moisture and acidity, as well as human activities such as agricultural and forestry management practices, affect soil biological communities and their functions. Soil biology is an interesting area of soil research and has yielded considerable information that is used in soil fertility management.

In vitro nematicidal activity of a terrestrial cyanobacterium, Synechococcus nidulans, towards plant-parasitic nematodes

The nematicidal activity of a terrestrial cyanobacterium, Synechococcus nidulans, was investigated. Extracts of S. nidulans cultures collected at weekly intervals for 5 weeks were sonicated and tested against second-stage juveniles (J2) of Meloidogyne incognita. Extracts of 2-week-old cultures caused the maximum immobility (94.2%) and mortality (29.3%) of J2, compared with controls (medium and water). This extract was tested in vitro against infective stages and hatch of M. graminicola, Heterodera cajani, H. avenae and Rotylenchulus reniformis. Extracts of sonicated S. nidulans caused a mean immobility in the range of 91.3-98.4% in infective stages of the nematodes, with no significant difference with an increase in exposure time from 24 to 72 h. The greatest mean percentage mortality was observed in M. graminicola (31.5%) followed by M. incognita (29.3%), H. avenae (20.9%), and R. reniformis and H. cajani (both 17.4%) with a significant increase with the period of exposure from 24 to 72 h. No significant differences in mortality were observed between M. graminicola and M. incognita and between H. avenae and H. cajani. The percentage hatch inhibition over control (water) was greatest in M. incognita (94.2%), followed by H. avenae (91.6%), H. cajani (72.3%) and M. graminicola (70.6%), and least in R. reniformis (58.6%).

Physiological studies on endorhizospheric establishment of Azotobacter chroococcum in wheat.

Ten strains of Azotobacter chroococcum were studied for their ability to invade the endorhizosphere of wheat. Strain W‐5 exhibited ability to invade endorhizosphere as shown in the microscopic observations. This strain was compared with the strain OA‐3 which did not invade the endorhizosphere zone. Strain W‐5 showed higher production of cellulase and pectinase than OA‐3. Both the strains induced defense enzymes in the host plant. However, induction of peroxidase and phenylalanine ammonia lyase activities (PAL) was higher in OA‐3 than W‐5. Quantitative differences in flavonoid like compounds obtained from root extracts and root exudates of plants inoculated with these strains were observed. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Formulation of a minimal nutritional medium for enhanced lipid productivity in Chlorella sp. and Botryococcus sp. using Response Surface Methodology

Chlorella sp. MCC 7 and Botryococcus sp. MCC 31 were investigated to enable large-scale biodiesel production from minimal constituents in the growth medium. Response surface methodology (RSM) was used to maximise the biomass productivity and lipid yield using only nitrogen (N), phosphorus (P) and potassium (K) as urea, single super phosphate and muriate of potash. The optimum values were 0.42 g/L nitrogen; 0.14 g/L phosphorus and 0.22 g/L potassium for Chlorella sp.; and 0.46 g/L; 0.14 g/L and 0.25 g/L for Botryococcus sp. Lipid yield of 42% for Chlorella sp. and 52% in Botryococcus sp. was observed. An enhancement in lipid yield by approximately 55% for Chlorella sp. and 73% for Botryococcus sp. was registered as compared to original nutrient medium. Fourier transform infrared (FTIR) analysis of extracted lipids revealed characteristic bands for triglycerides. This study provided utilisation of a practicable nutrient recipe in the form of N, P, K input for enhanced lipid yield from the selected microalgal strains.