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Being creative and resourceful in the biofuels industry

Throughout history, fossil fuels were the dominant energy source that powered the world.
However, due to their negative environmental impact and unsustainability, there have been new efforts to shift the world’s reliance onto greener, renewable sources of energy, such as biofuels. The biofuel industry has become an emerging market, providing a multitude of alternative energy sources that can one day power everyone’s lives. Interestingly, different parts of the world are tackling biofuel production in creative and resourceful ways.
There have been recent developments in South Africa, Brazil, Canada, and Oman that have all found effective and unique methods to produce biofuel.
In South Africa, researchers have examined fungi from the manure of locally wild herbivores to determine their usefulness in bioethanol production. Previous reports have analysed fungi from the manure of domestic animals, but few have pertained to wild herbivores.
Normal methods of processing lignocellulose into bioethanol include pretreatment, hydrolysis, fermentation, and distillation. The manure of herbivores is a natural source of pretreated lignocellulose, which becomes a hotbed for fungi [1].
Fungi cultivated from this manure can then be investigated for their ability to either create enzymes to aid in hydrolysis or to ferment xylose into ethanol, which are the main purposes of their research.
The methodology behind this research takes advantage of resources local to South Africa.
The fifty manure samples analyzed came from eight wild herbivores living in Kruger National Park in South Africa [1]. To isolate the fungi on the manure, the researchers sprinkled small amounts of the samples onto agar plates to extract pure cultures of the fungi. In total, 36 yeasts and 65 molds were isolated.

The extracted yeast isolates were tested for their ability to ferment xylose. Those that were successful in fermentation were then tested for their viability for growing at higher temperatures and in the presence of acetic acid and furfural, which are known inhibitors present in the pretreatment process.
The mold isolates were analyzed, using a thatch grass-based medium, for their enzyme production ability relating to endoglucanase, xylanase, and mannanase [1].
Six of the 36 yeast samples successfully produced ethanol while growing on xylose [1]. One yeast developed from dassie manure, known as Pichia kudriavzevii KP34ey, had the highest concentration of ethanol on xylose at 2.3 ± .03 g/L [1].
This concentration is comparatively lower than those from glucose fermentation, but further investigation into this yeast with a more favorable substrate may yield stronger results [1]. This yeast showed strong resistance to inhibitors, as it exhibited the highest tolerances to the presence of acetic acid (3 g/L), furfural (1 g/L), and high temperatures [1].
The removal of these inhibitors beforehand would make ethanol production more expensive, making ethanol production using Pichia kudriavzevii KP34ey more cost-effective. On a small scale, this yeast shows strong potential, making it a good candidate for future examination to determine its commercial viability. Concerning the mold isolates, the best enzymatic strains were produced from three particular molds— Aspergillus sp. KP29 mm, Aspergillus brasiliensis KP39 nm, and Aspergillus sp. KP35 mm [1].
The results were not surprising as the Aspergillus species had been previously documented for its ability to produce plant polysaccharide degrading enzymes [1]. With further study into the fungi that this project highlighted, perhaps the capabilities these fungi have can be fully realized in bioethanol production.
When tackling the problem of producing biodiesel from waste, researchers in Brazil opted for a different solution by producing biodiesel from waste vegetable oil using catalysts developed by extracting lithium from discarded lithium-ion batteries.
Minimising feedstock costs is a common strategy used to make biodiesel production more economically viable, making waste cooking oil (WCO) an attractive resource for this purpose. One technique used to convert pretreated WCO into biodiesel is alkaline catalysis, where sodium hydroxide and potassium hydroxide are commonly used catalysts to aid in transesterification [2]. The researchers in Brazil have modified this process by adding a key ingredient— lithium.
Their strategy behind this innovative use of lithium is to minimize the waste and negative environmental impact that results from discarded Li-ion batteries while making the biodiesel production process cheaper and more viable [3].
The methodology behind this investigation begins with how the materials were sourced. The WCO was sourced from local establishments and residences in the city of Vitória, Brazil [3]. They cultivated the lithium from discarded Li-ion batteries for the metal hydroxide mixture used for catalysis [4].
The complete methoxide mixture included mixes of LiOH + NaOH and LiOH + KOH with methanol [3]. The researchers mixed the WCO sample into the methoxide catalyst system until the transesterification reaction went to completion [3]. Subsequently, the biodiesel and glycerol were separated using a separation funnel and rinsed to remove the catalyst excess [3]. The biodiesel was further characterized using tests such as thin-layer chromatography and hydrogen nuclear magnetic resonance spectroscopy [3].
The results successfully show the potential of lithium being used in this process. Good separation between the biodiesel and glycerol in the separation funnels showed the completion of the transesterification reaction. In addition, the chromatograms comparing biodiesel with WCO reveal the purity of the biodiesel.
Using the 1H NMR spectra procured for biodiesel made from the LiOH + NaOH mixture and the LiOH + KOH mixture, the calculated conversion yield was 90% and 89%, respectively [3]. These are promising yields and are considered to be relatively high. A future investigation regarding this process is the effect that lithium has on biodiesel as a contaminant, which is an important attribute to know for large-scale commercial production using this process. Overall, these results can have widespread implications on the biofuel industry and environmental impact as lithium sourced from waste lithium-ion batteries prove to be an effective aid in catalysis for biodiesel production.
Researchers in Canada at the Institut National de la Recherche Scientifique (INRS) are finding inventive ways to make biodiesel production cheaper by sourcing carbon from renewable wastes like crude glycerol and municipal sludge and using a novel method for lipid extraction using a biodegradable surfactant.
The challenge these researchers set out to resolve is the high cost of the biodiesel production process as it relates to using microbial oil sources and glycerol byproduct. The current production cost of using a microbial oil source is at an expensive $5.9/kg biodiesel, compared to the commercial biodiesel price of $1.38/kg biodiesel [5]. To combat this cost, the Canadian researchers are using municipal sludge as an economic measure to foster the growth of oleaginous microorganisms [6]. Another cost issue relates to the expensive purification process of crude glycerol byproduct from typical biodiesel production [7].
An established use for crude glycerol is bioconversion, which is why the researchers are using it for its substantial source of carbon as a fortifying substrate to the municipal sludge for microbial oil production [8,9].
The INRS process begins with inoculum development and production fermenter, where they cultivated Trichosporon oleaginosus, the microbial strain used for lipid production. The municipal sludge went through a pretreatment process, which subsequently had sterilized crude glycerol added to it as a secondary carbon source [6,8]. The researchers developed a novel method for biomass settling using extracellular polymeric substances (EPS) as the bio-flocculant and calcium chloride as a chemical coagulant [8].
These substances helped to provide the biomass with a surface charge to aid in binding with other suspended particles for floc formation [10]. For the lipid extraction and recovery, the researchers created an alternative method using bio-surfactant and free-nitrous acid to further cut costs to production. Subsequently, the lipids were recovered by using petroleum diesel to contain them [11]. In the final step, the petroleum diesel with the dissolved lipids was reacted with methanol for the transesterification reaction to create the final biodiesel (type B10), which was separated from the glycerol byproduct in a phase separation tank [8].
After price calculations, the cost to produce biodiesel using their novel process is $0.72/L, which is much less than the $6.78/L from conventional microbial oil sources [8]. The production costs compared well to biodiesel costs produced by using vegetable oils ($1.15/L) and waste cooking oils ($0.85/L) [5,12].
At a commercial level, these prices are still higher than they need to be to make sense financially, but the results lay the groundwork for future development of this concept. Another beneficial result of this process was the reduction of greenhouse gas emissions from repurposing the municipal sludge, as the net capture of CO2 in the process is 10.78-ton CO2 eq./ton B10 biodiesel [8]. Overall, this project made great headway in proving the potential economic viability of this novel process and minimizing waste by using crude glycerol and municipal sludge.
Researchers in Oman went with a different approach to producing biodiesel by making the source of feedstock and catalyst to be the same.
Omani researchers are using waste date pits as a renewable heterogeneous catalyst and as an oil feedstock to be used for biodiesel production. Date palm is the leading crop grown in Oman, taking up 54% of the agricultural land of the country, making waste date pits a cheap, inedible biomass source that is an excellent alternative source for producing biodiesel [13,14]. Using these date pits to create a heterogeneous catalyst simplifies the separation process between the catalyst and the reaction mixture.
The procedure to their investigation starts with the locally sourced waste date pits, which were collected from an industrial business in Muscat, Oman [14]. Once date pit oil was extracted using a Soxhlet extractor, the remaining date pit powder used for the synthesis of the catalyst was prepared and saturated with an aqueous solution of calcium nitrate tetrahydrate at 2%, 4%, and 6% of calcium [14].
The resulting products were then calcined in a furnace to create the final catalysts termed as C, C1, C2, and C3 for 0%, 2%, 4%, and 6% of calcium in the carbon, respectively [14]. These catalysts were then characterized for a multitude of properties. For the biodiesel production process, the date pit oil was first heated before adding methanol and synthesized catalysts [14]. Once the transesterification reaction went to completion, the product was cooled and centrifuged to remove the solid catalyst [14]. The biodiesel was then recovered using a separation funnel to separate the biodiesel from the glycerol byproduct and was purified [14].
The catalyst that performed the best was C2 with a biodiesel yield of 98.2%, which is a very high result [20]. In addition, the quality of the biodiesel met the standards set by both ASTM and EN regulating agencies, proving its suitability to be used as a fuel [14]. The researchers could produce 100 mL of pure biodiesel from 10-15 kg of waste date pits and had less harmful emissions than fossil fuel emissions and a lower greenhouse gas footprint [15].
The catalyst was proven to be reusable, as it showed no significant decrease until after the sixth consecutive test [14].
The fundamental difficulty in applying this concept for commercial purposes is the scalability of the process. Currently, it would not be feasible to conduct large scale operations using only date pits for the amounts of date pits that would be required to do so would be tremendous. However, the goal of these researchers was to reduce the dependency on fossil fuels by creating a diverse choice of alternative energy sources, in which this project is a step in the right direction [15]. Nevertheless, the novel catalyst produced by using waste date pits has proven to be effective in biodiesel production and provides yet another green alternative to using fossil fuels.
These examples demonstrate a worldwide effort to minimize our reliance on fossil fuels and shift our dependencies to green, renewable, and sustainable sources of energy. Biofuel production is an integral part of that goal.
With a diverse choice of methodologies on how to produce biofuel based on the local resources available in a particular region, there are solutions for every part of the world. The innovative methods that researchers have developed globally are remarkable, and their work will continue to lead our world into a cleaner future. As we continue to investigate innovations in this field, perhaps some of these breakthroughs can migrate from the small lab scale setting in which they were conceived to a broader commercial application. Whether it be developing yeasts from manure, using Li-ion battery waste or waste date pits for catalysis, or growing microbial oil on municipal sludge, these are all worthwhile endeavors to further advance the production of biofuels.

Dr. Raj Shah is a director at Koehler Instrument company, NY, an adjunct professor of chemical engineering at State university of NY, and is a Fellow both at the Royal society of Chemistry and the Chartered management Institute, London
Mr. Nabill Huq is a student of Chemical engineering at SUNY, Stony Brook, where Shah is the chair for the external advisory board of directors

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