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2020 Annual Report
Objectives
1: Enable commercial processes for transforming animal protein into new marketable flocculants.
1a. Develop processing techniques for the solubilization of rendered protein with emphasis on intact proteins and high molar mass protein fragments.
1b. Evaluate technological alternatives for transforming raw chicken blood into a high potency flocculant at low processing cost.
2: Enable market growth for flocculants based on animal proteins by improving their performance and expanding their market applications.
2a. Apply a series of covalent modification strategies to improve blood and rendered protein flocculant performance.
2b. Identify particular application areas to which blood and rendered protein flocculants are well suited.
Approach
Both rendered protein and chicken blood have inherent flocculant properties, but these substances also have other properties which make them unsuitable for commercial flocculant applications in their ‘raw’ state. Poor solubility is a primary obstacle to commercial utilization of rendered protein as a flocculant. Instability, high water content, and dark red color are among the obstacles to blood utilization. The project will focus on developing processing techniques for surmounting these obstacles under Objective 1. With the current state-of-the-art, rendered protein or blood flocculants have significant performance limitations. Improving their performance through covalent modifications is the focus of Objective 2a. Finally, any class of flocculants is well suited to some particular application areas and not to other areas. In Objective 2b, the focus is on identifying particular application areas appropriate for rendered protein and blood flocculants.
Progress Report
Rendered protein meals, such as meat & bone meal (MBM), may be good sources for flocculant proteins, but MBM proteins have poor solubility. Past attempts at solubilizing this protein produced unsatisfactory results; either the proportion of the protein solubilized was too low or the protein was digested into fragments that were too small to be useful. This project’s Objective 1a explored new approaches to this problem. We adapted an obsolete method found in old protein structural analysis literature. The method uses dilute acid and carefully selected conditions to cut proteins at very specific sites; this allows the protein to be solubilized without having it digested into very small fragments. Although the method had not previously been used with substrates like MBM, our results show that it works well. The solubilized protein exhibits better flocculant activity than had previously been achieved using hydrolyzed MBM protein. Further work along these lines adapted the method to the practical constraints of an industrial process. It was found that two processing steps could be eliminated entirely, and that the reaction could be run at a much higher substrate concentration as long as the acid concentration was increased to compensate. A second approach to MBM solubilization was investigated which involved breaking protein crosslinks, but high processing costs and limited efficacy led abandonment of this line of research.
Our work in past projects has shown that blood or blood components can function as very effective flocculants. Objective 1b was intended to examine how a blood-based flocculant could be commercially produced. Although past work has suggested that the economics of a blood-based flocculant product are favorable, we recently examined this in much greater detail by conducting a techno-economic analysis of a hypothetical commercial-scale facility processing blood into a flocculant product. The analysis included details beginning with collection and transport of raw blood to the packaging of the final product. The results suggest that such a facility could operate profitably and market a product that would be price competitive with the conventional flocculant known as PAM.
Hemoglobin is a component of blood that is responsible for most of the flocculating activity of blood. Until now, it has not been clear whether the extra expense of isolating hemoglobin from blood is justified for the purposes of producing a flocculant product. Our recent research into this issue has shown that hemoglobin isolation provides only a small benefit of increasing the flocculant potency. Whole blood was expected to add more pollutants to water than isolated hemoglobin, but this was shown not to be the case when the flocculant is used at appropriate doses. The previously mentioned techno-economic assumed that hemoglobin isolation was required, so the finding that this is not required suggests the processing costs could be greatly reduced by omitting certain steps in the processing. Finally, we developed improvements to the process of washing, concentrating, and drying the product of a protein modification reaction. Making the product in a membrane-concentrated liquid form has been proven superior to more expensive dehydration by freeze-drying, and the resultant product fits more readily into end users’ existing liquid dosing systems.
While proteins can be very effective flocculants, they do suffer from certain limitations. Objective 2a sought to overcome these limitations by applying chemical modifications to protein, resulting in products that are more attractive for technology transfer. One particular limitation is diminishing effectiveness when the acidity of the water to be treated is too low. To mitigate this limitation, a simple chemical reaction was used to modify protein; the modification was anticipated to improve the flocculant performance of protein under low acid conditions. Experiments compared normal and modified protein in terms how well they clarify water, and the dose required to achieve good clarification. The results show that the modification had the desired effect of improving performance in water that is near neutral pH with three out of the four animal proteins tested. The modification also had the unexpected effect of significantly improving overall flocculant performance; under some conditions, the modified hemoglobin could clarify water as well as unmodified protein with only one quarter the dose; another protein which normally has very little flocculant activity became a fairly potent flocculant when modified. Further research provided a deeper understanding of the effects of modification at a molecular level. Overall, this research has provided a simple, inexpensive way to improve the performance of a protein-based flocculant. Such an improvement will expand the opportunities for commercial application of the flocculant.
Under the same objective, improved performance was also achieved through a strategy of increasing the size of the flocculant molecules. Hemoglobin and albumin from slaughterhouse blood were successfully cross-linked by chemical or enzymatic means. If the crosslinked was appropriately controlled, the resulting products were much larger than naturally occurring proteins and had the anticipated increase in performance. The same cross-linking approach was applied to the solubilized MBM protein produced under Objective 1a, but in this case the treatment was less successful and the performance gains were modest. Modification of proteins as described earlier followed by cross-linking had a synergistic effect on performance, resulting in some of the highest performance yet observed, although such products would have a high production cost.
Finally, Objective 2b focused on identifying particular application areas to which blood and rendered protein flocculants are well suited. Blood-based flocculants were used to treat an ethanol plant by-product stream known as “thin stillage.” Normally, the solids and nutrients in thin stillage are recovered by evaporating the water, which requires a large amount of energy. Recovery of solids by flocculation is much less energy intensive. Research showed that pig blood or purified hemoglobin could effectively recover the suspended solids in thin stillage. This flocculation treatment by itself did not recover phosphate from the stillage, but it was found that it could be used in conjunction with another treatment that causes phosphate to precipitate.
Under a CRADA with a Wisconsin based firm, blood-based flocculant product was adapted to the recovery of nutrients and solids from liquid manure. This collaboration expanded the scope of the technology, because rather than flocculation, the CRADA partner uses a system in which rising air bubbles carry contaminants away from the clean liquid; the products performed well in this type of system. Collaboration under a separate agreement has evaluated the ability of protein flocculants to serve as conditioners in the processing of a wastewater treatment by-product; the results to date have been promising.
Finally, a series of projects was completed on a topic that was not part of the planned objectives. Hemoglobin was used to produce an insoluble substance that can be used like activated charcoal to remove organic pollutants from water. This substance has the added benefits of being magnetic, so that it is easily recovered, and being re-usable after the adsorbed pollutants are removed. This substance was found to be particularly useful for removing dye contamination from industrial wastewater. This technology is expected to create a new use for slaughterhouse blood, and has been the subject of two invention disclosures.
Accomplishments
Review Publications
Essandoh, M., Garcia, R.A., Gayle, M.R., Nieman, C.M. 2020. Performance and mechanism of polypetidylated hemoglobin (Hb)/iron oxide magnetic composites for enhanced dye removal. Chemosphere. 247:1-9. https://doi.org/10.1016/j.chemosphere.2020.125897.
Essandoh, M., Garcia, R.A., Neiman, C., Strahan, G.D. 2020. Influence of methylation on the effectiveness of meat and bone meal protein as a bioflocculant. Journal of Agricultural and Food Chemistry. 122:55-61. https://doi.org/10.1016/j.fbp.2020.03.009.
