As the lubricant industry moves toward sustainability, research efforts are underway to determine how to use naturally derived products in the formation of hydrocarbons that can be used in fuel, as base oils and as precursors to polyalphaolefins (PAOs). The challenge in undertaking this approach is how to efficiently remove the oxygen-containing components in naturally derived products that are typically ester and alcohol derivatives.

In a previous TLT article,¹ a potentially sustainable pathway to produce base oils from plastic resins was discussed. Plastics have come under scrutiny for polluting the environment leading the industry to look for ways to reuse them. Researchers determined that a mesoporous silica-based heterogeneous catalyst can convert high-density polyethylene to hydrocarbons with chain lengths between C9 and C35 with a peak at C14. The pore size of the catalyst appears to have a substantial effect on the carbon chain distribution.

Zhen Wang, assistant professor in the department of biological sciences at the University of Buffalo in Buffalo, N.Y., says, “Development of a pathway to produce hydrocarbons from naturally derived products in a living microbe involves identifying the right type of enzymes to facilitate the sequence of steps needed to build a hydrocarbon structure and then remove the oxygen through a decarboxylation-dehydration process. The main enzymes available to manufacture hydrocarbons naturally are situated in the cellular membranes. These enzymes need other enzymes to act as reducing partners to eliminate oxygen, which is of low catalytic efficiency. The presence of these enzymes in the cell membrane also means that their abundance in the engineering microbe will be limited.”

In identifying an approach to produce hydrocarbons, Wang and her colleagues recognized that an ideal strategy would be to have microbes synthesize medium-chain 3-hydroxy carboxylic acids through a chain elongation starting with acetyl-coenzyme A (CoA), which carries the two carbon acetyl group. She says, “3-hydroxy carboxylic acids contain the proper functionality to undergo decarboxylation, which leads to the removal of oxygen from the hydrocarbon chain. In addition, the process of manufacturing this organic derivative does not require the use of reducing agents.

A second concern for the researchers was to identify enzymes that will not require the use of the cellular energy source, ATP. Wang says, “Cells that participate in the fatty acid chain elongation will utilize two ATP molecules per two carbon elongation.”

The researchers have developed a new pathway to 3-hydroxy carboxylic acids that does not rely on ATP and the enzymes used and does not interfere with the cellular fatty acid biosynthesis.

Thiolases
Wang and her colleagues identified a group of enzymes known as thiolases as potential candidates for use in chain elongation. She says, “The currently used thiolases are not selective for the synthesis of medium-chain alcohols and carboxylic acids. In evaluating an enzyme cluster present in mycobacteria, we found that PaaJ-like thiolases have the potential to catalyze the carbon chain elongation.”

The starting material used by the researchers is the sugar, glucose, which can be converted enzymatically to acetyl-CoA through glycolysis, which is native to the microbe. Thiolases are able to produce medium-chain acyl-CoAs through a series of non-decarboxylative Claisen condensations.

Solubility issues with the thiolase identified from a specific mycobacterium led the researchers to search a different phylum of bacteria known as proteobacteria. Wang says, “We found nine thiolases present in proteobacteria that could potentially be effective in carbon elongation. Experimental analysis of these thiolases led to the selection of three candidates that are able to produce C8-C10 acyl-CoA, in vitro.”

The researchers then placed genes encoding the enzymes into a microbial host which is a strain of the bacterium E. coli, added nutrients and shook them in an incubator under fermentation conditions to promote the carbon elongation process. The main products obtained are C8 and C10, 3-hydroxy carboxylic acids. Figure 1 shows a series of flasks used in these experiments.


Figure 1. Incubating flasks containing glucose and three thiolase enzymes are shown. They produced C8 and C10, 3-hydroxy carboxylic acid intermediates which are then transformed into C7 and C9 alpha olefins. Figure courtesy of the University of Buffalo.

To complete the process of producing alpha olefins, the researchers determined that use of heterogeneous catalysts was needed to do the decarboxylation and dehydration steps. While it would be desirable to use enzymes in these processes, the researchers found that there were limitations including high ATP expenditure, additional reductases and enzyme solubility issues. 

The researchers decided to work with the Lewis acid catalyst, niobium pentoxide, which reacted with microbial products extracted from the fermentation broth to produce C7 and C9 alpha olefins. Wang says, “Future work will focus on determining how the overall product yield for the conversion from glucose to alpha olefins can be improved.”

This study serves as a proof of concept that the combination of an enzymatic carbon elongation process followed by heterogeneous catalysis can convert the natural sugar, glucose, to alpha olefins that can fundamentally be used to produce PAOs. Wang believes this approach can be used to produce both linear and branched hydrocarbons. Using enzymes as the catalysts should enable the preparation of specific hydrocarbons with a low concentration of byproducts.

Additional information on this work can be found in a recent article² or by contacting Wang at zhenw@buffalo.edu or her co-author, Michelle Chang, professor of chemistry at the University of California-Berkeley (mcchang@berkeley.edu).

REFERENCES
1. Canter, N. (2021), “Preparation of base oils from plastic resins,” TLT, 77 (3), pp. 16-17. Available here.
2. Wang, Z., Song, H., Koleski, E., Hara, N., Park, D., Kumar, G., Min, Y., Dauenhauer, P. and Chang, M. (2021), “A dual cellular-heterogeneous catalyst strategy for the production of olefins from glucose,” Nature Chemistry, 13 (12), pp. 1178-1185.