Reducing Cost of Dewatering/Harvesting

Algae Update 5

TLDR:

There are 3 main cost categories for algae. They are cultivation, dewatering, and drying. In my previous update, I worked on decreasing the cost of cultivation. This week, I will go through my current hypothesis to decrease the cost of harvesting/dewatering. Note: Harvesting and dewatering refer to the same action of separating algae from the water.

What is Dewatering?

Separate algae from the water. Once the algae are separated from the water, aggregates of wet algae biomass can be formed. You would normally take this wet biomass and dry it to maximize your density of algae biomass (to be discussed next update).

Dewatering is about 30% of total algae cost. When you put in mind that the status quo cost of algae biomass is $5/kg, which means your dewatering cost is about $1.5/kg.

Status Quo of Dewatering

There are many different ways to dewater algae including physical, electrical, and chemical methods.

Physical methods include centrifugation, filtration, and flotation. They can achieve high efficiencies, but there is a significant energy requirement. This increases the cost of algae biomass where it is not feasible.

Gravity sedimentation can greatly save energy during operation, but the time consumption and species-specific feature limit its wide application.

Negatively charged algae can also be concentrated by the electrical method, whereas the establishment of an electric field requires massive capital expenditure. The chemical method that is involved with the utilization of organic and inorganic flocculants can aggregate microalgal cells in a short time, but the complicated reagents may result in biomass contamination and the decrease of biofuel values.

In this update, I will be investigating fungi as a potential flocculant for cost-effective dewatering with algae. This hypothesis initially came from the understanding that fungi and algae have co-existed together for the last 500 million years. Together they are described as lichen, which refers to the symbiotic relationship between algae and fungi. Fungi secret extracellular enzymes which can convert solid organic matters into soluble nutrients and carbon dioxide, making it easier for microalgae cells to be assimilated. In turn, microalgae release oxygen (through photosynthesis) to promote fungal respiration.

Introduction to Filamentous Fungi for Harvesting

Filamentous fungi can immobilize microalgae through mycelial (root-like structure of fungi) interactions. Immobilization refers to preventing algae from attaching to water. When algae are not attached to water, the fungi will help algae aggregate. The biomass output can later by a sieve (wire or plastic mesh) because of their relatively large size (>1 mm in average diameter). This process takes no energy and your only input cost is buying fungi.

There are two types of co-cultivation modes for harvesting: fungal spore-assisted or the pellet-assisted method. Fungal spore-assisted harvesting is the process of co-culturing fungal spores (one-celled unit that multiplies through asexual reproduction) organisms with microalgae to form aggregations. Forming an aggregation is creating a cluster of algae biomass. The pellet-assisted method happens by adding pre-cultured fungal pellets directly.

The Outcome of Harvesting With Fungi

Based on this review paper, out of 15 pellet-assisted harvesting experiments, the harvesting efficiency is between 65% and 99% with timing between 0.5 hours and 72 hours. There were also 10 spore-assisted harvesting experiments which included a harvesting efficiency between 93% and 100% with timing between 4 hours and 72 hours. The difference in harvesting efficiency is dependent on algal and fungal strain interaction since harvesting efficiency is species-specific. When you change the algae strain or fungi strain, you would get different results.

The key difference between both methods is most pellet-assisted experiments tend to have a timeline under 24 hours whereas spore-assisted tend to have a timeline between 24-72 hours. The majority of pellet and spore assisted tends to be above 95% efficiency. If there is a difference in efficiency from either method it would mostly be a matter of diminishing returns. Regardless of the general tendencies, you also have outlier combinations. For example, Chlorella Vulgaris (algae strain) combined with Aspergillus sp. (fungal strain) was able to get >97% harvesting efficiency in under 4 hours with the spore-assisted method.

The outcome of high harvesting efficiency comes from 3 major interactions. The main interactions are electrostatic, hydrophobic interactions, and specific interactions with components of the cell walls.

Electrostatic Interaction

Depending on the pH value in the culture system, ionizable groups which are molecules that can be broken down into ions (elements that have a charge) can be protonated (add a proton/increase positive charge) and deprotonated (losing a proton/increasing negative charge) to create charges on the surfaces of algal cells.

The alkaline pH (pHs greater than 7) of microalgal culturing conditions the ionizable groups of algae cell walls deprotonate to be negatively charged. The net negative charges on cell walls induce electrostatic repulsion between algal cells, pushing them away from each other.

The fungal spores or pellets are introduced to the algae culture medium to stop suspension in water and create aggregations of algae.

For the pellet-assisted method, since the pH value in the fungal medium (where fungi are cultured before being introduced to algae culture) is acidic, the surface functional (carboxylic and amine) groups of mycelium remain protonated, leading to the net positive charges of the fungal hyphae. Therefore, when positively charged fungi contact negatively charged algal cells, charge neutralization can fully display advantages to eliminate the Zeta potential. Zeta potential is an efficient indicator to measure the degree of electric force which avoids aggregation of particles. Once the systematic Zeta potential is near zero, the repulsive cells can approach each other by the attraction of charge, hydrogen bonding, and van der Waals forces, subsequently occurring co-pelletization.

For the spore-assisted method, the spores are introduced to an algae culture. Due to their natural acidic pH (less than 7), they change the pH of the system. The fluctuation of ambient pH alters the charged properties of cell walls inducing electrostatic attraction.

The difference between both methods is mainly time. Fungi by default have a pH of less than 7 (spore), when it is cultured (grown before adding to algae culture) you make the fungi have lower pH to increase the force of the positive charge to speed up the algae and fungi attraction, increasing harvesting time. Most pellet-assisted experiments tend to take 24 hours whereas spore-assisted tend to take between 24-72 hours.

Hydrophobic Interaction

Hydrophobicity is the property of an organism that repels water.

The hydrophobicity of filamentous fungi is generated by surface-active proteins, known as hydrophobins. Hydrophobins are a group of small (~100 amino acids) proteins that are expressed only by filamentous fungi. They are known for their ability to form a hydrophobic (water-repellent) coating on the surface of an object (this would be algae).

The initial electrostatic attraction of both fungi and algae would create aggregation and the water-repelling property would create another force to have algae separate from water (once connected to fungi).

Interactions with Specific Components on Cell Wall of Fungi

Talukder et al. found a positive relationship between the content of chitin (the primary component of cells walls in fungi) and microalgal immobilization (or harvesting) efficiency because the strongly charged chitin could function as a cationic (or a positively charged) flocculant to neutralize the negative charges on the surface of algal cells.

Similarly, the amorphous polysaccharides on the cell walls of fungi, usually bonded with proteins as glycoproteins, also play an important role in cementing other microbial cells by delivering positive charges, helping attraction of algae and fungi.

When certain divalent ions (an ion with 2+ or 2- charge) exist in the solution, the components on fungal cell walls can be linked with microalgal cells to form aggregations through ions bridging. Li et al. measured the calcium-binding ability of Streptomyces sp. pellets, and the final results suggested the addition of 5 mM (millimolar) of CaCl2 (an ion with a 2+ charge present in fungi) displayed the highest flocculation efficiency.

Li et al. investigated the critical function of proteins on fungal mycelium (root-like structure of fungi) during the flocculation (aggregation) process. When proteinase K (found in extracts of Engyodontium, a genus of fungi) was added into microalgal suspension with fungal pellets. The structure of proteins on the cell walls of the fungi was destroyed and the flocculation efficiency was only 0.11 times in comparison with the control without any addition.

These are a few of the many mechanisms within the fungi cell wall to help increase or decrease harvesting efficiency.

Main Variables That Determine Harvesting Efficiency (In Order of Importance)

  1. Fungal-algal strain + type of fungi harvesting — They determine the main interactions that occur. When you change the algae strain or fungi strain, you would get different results.

    Algae strain — Chlorella Vulgaris (since it has the highest CO2 capture rate)

    Fungi strain — to be determined. The current placeholder (based on 1 paper) is Aspergillus sp. with Chlorella Vulgaris created a >97% harvesting efficiency in 4 hours. This happens with using pretreated molasses (or distillery/liquor production) wastewater.

    Since I’m already thinking about using Chlorella Vulgaris with some type of wastewater, this would be a good place to start (for experiments).

    Type of fungi harvesting method — to be determined. The example mentioned above was a spore-assisted harvesting method.

  2. Nutrients source — Glucose

    When referring to the nutrient source, I’m not talking about nitrogen and phosphorus (the main nutrients for algae). These are nutrients specifically to enable fungi growth. Since fungi are heterotrophs they rely on eating other organisms for growing. One of the inputs they need is a source of carbon. This can be glucose, glycerol, and acetate. All have different effects so they should be tested for. In general, glucose is the most suitable organic carbon source for fungi and algae to be utilized, and the concentration ranges from 2 to 20 g/L in diverse conditions.

  3. Fungi Algae ratio — fungal spores and algae during the growth period may compete for limited resources in the medium due to the inappropriate inoculation ratio, which results in the excessive predominance of one species. A high concentration of spores means that more available nutrients in the medium need to be assimilated by fungi to support metabolism, forming a larger size of pellet to capture algae into the mycelium and further inhibiting the growth and biomass productivity of algal cells.

    For example, the ratio of Aspergillus fumigatus (fungi strain): Botryococcus braunii Kossou-4 (algae strain) with the range from 1:20 to 1:50 has decreased the harvesting efficiency from 97% to 35%.

  4. Temperature — 20° to 30°C

    Here you are optimizing for the optimal temperature for both algae and fungi. It will be in the range of 20-30 degrees (since that is optimal for algae). When you go higher than 30 degrees the algae starts to deteriorate which would not be helpful for harvesting or the biomass output. This will likely involve the selection of fungi strains specifically to be under 30**°**C.

  5. pH

    Most papers suggest between a 7.0 to 8.0 pH for algae so you would specifically find fungi strains to fit this range (similar to how it would be done with temperature)

What Does Fungi Harvesting Mean For Algae Cost?

Based on the last update. The status quo cost breakdown would be 50% for cultivation, 30% for dewatering, and 20% for drying. Cultivation would be $2.5/kg, dewatering would be $1.5/kg, and drying would be $1/kg.

I brought down cultivation cost to $0.12. With fungi as the harvesting tool, the only input cost is the initial fungi itself, similar to the cost of algae cells they are negligible to the point where they are not accounted for in operating cost. In this case, we can assume that all $1.5/kg for dewatering is gone. Now you only have cultivation cost and drying cost getting you to $1.12/kg of algae. Keep in mind there is probably some margin of error about the current cost I’m proposing vs when I test it.