Nylon 6 is widely used in high-performance materials. However, the inherent structural rigidity of nylon 6 inhibits catalytic depolymerization to its monomer, ε-caprolactam. Here, we demonstrate an acid-catalyzed alcoholysis process that depolymerizes nylon 6 to ε-caprolactam, achieving yields up to 74% using n-propanol as solvent and phosphoric acid as catalyst at 220°C for 2 h. We further applied the process to commercial nylon-containing products, demonstrating ε-caprolactam yields of 67% to 76%. Process modeling and techno-economic analysis estimated a minimum selling price of recycled nylon 6 at $1.79/kg, 30% lower than the 5-year average market price of virgin nylon 6. These estimates depend on assumed costs and process performance and may vary with future conditions. Life cycle assessment indicated that nylon 6 produced via alcoholysis can reduce greenhouse gas emissions by up to 63% compared with primary production. Overall, this study demonstrates a facile, cost-effective, and environmentally beneficial process for nylon 6 chemical recycling.

Viable manufacturing pathways to produce bio-based chemicals from renewable feedstocks, such as lignin derived from plant biomass, are needed to decarbonize the chemicals manufacturing sector. Converting the recalcitrant lignin polymer to valuable bioproducts remains a longstanding challenge in biorefining, with the highest reported single-product yield from lignin currently around 20 wt% (refs. 1-4). Most existing lignin depolymerization strategies target aryl-ether bond cleavage, which can produce aromatic monomers in yields of only about 30 wt%, and still as complex mixtures with C-C-linked dimers and oligomers5,6. The recalcitrance of these C-C linkages between aromatic moieties fundamentally limits single-product yields from lignin, prompting the development of strategies to efficiently cleave these C-C bonds3,7-9. Here we show how reductive processing of lignin from poplar accesses a hydrocarbon mixture of alkyl-aromatic monomers and oligomers that is privileged for oxidative conversion to monomeric aromatic carboxylic acids, comprising mostly benzoic acid and phthalic acid isomers in up to 73 wt% monomer yields, using a Co/Mn/Br catalyst. The soil bacterium Pseudomonas putida KT2440 was engineered to convert this mixture of aromatic carboxylic acids to muconolactone, a precursor to bio-based nylons, enabling final adipic acid yields up to 26 wt% (gram adipic acid per gram lignin) with a maximum theoretical yield of 57 wt%. This pairing of reductive and oxidative steps with lignin resembles processes in petrochemical refining and shows how lignin may be converted into a single, valuable bioproduct in high yields.

Polyhydroxyalkanoates (PHAs) are versatile, biobased polyesters that are often targeted for use as degradable thermoplastic replacements for polyolefins. Given the substantial chemical diversity of PHA, their potential as cross-linked polymers could also enable similar platforms for reversible, degradable thermosets. In this work, we genetically engineered Pseudomonas putida KT2440 to synthesize poly(3-hydroxybutyrate-co-3-hydroxyundecenoate) (PHBU), which contains both 3-hydroxybutyrate and unsaturated 3-hydroxyundecenoate components. To reduce the brittleness of this polymer, we physically blended PHBU with the soft copolymer poly(3-hydroxydecanonate-co-3-hydroxyundecenoate) in mass ratios of 1:3, 1:1, and 3:1. Upon observing varying degrees of immiscibility by scanning electron microscopy, we installed dynamic boronic ester cross-links via thiol-ene click chemistry, which resulted in compatibilized dynamic thermoset blends ranging in hard, medium, and soft rubber or elastomer thermomechanical profiles. These dynamic thermoset blends were subjected to controlled biological degradation experiments in freshwater conditions, achieving timely mass loss despite the cross-linked architectures. Overall, this work highlights a two-component platform for the production of degradable and reprocessable dynamic thermoset blends suitable for several classes of cross-linked polymer technologies from tailored, biological PHA copolymers.

Lignin-derived aromatic carboxylic acids can be produced from oxidative catalytic processes and are promising building blocks for performance-advantaged bioproducts that leverage their inherent heteroatom functionalities. Here, we synthesize 2-methoxyterephthalate and 2,6-dimethoxyterephthalate derivatives by electrochemical carboxylation of guaiacyl- and syringyl-derived lignin monomers obtained from the oxidative deconstruction of lignin. These methoxylated terephthalates are evaluated as co monomers in poly(ethylene terephthalate) (PET) and as plasticizers that could replace petrochemically derived isophthalate and phthalate, respectively. Specifically, we co-polymerize 2-methoxy- and 2,6 dimethoxyterephthalate with dimethyl terephthalate to form several PET co-polymers, both of which enable the properties of PET to be tuned, with an incorporation beyond 25% producing amorphous polyesters. At 10 mol% loading in the co-polymers, we demonstrate that the bio-derived co-monomers exhibit comparable behavior to isophthalic acid, a commonly used co-monomer in PET, by lowering the crystallinity and melting temperature. Moreover, methoxyterephthalate esters (2-ethyl hexyl and butyl) are compared to phthalate and terephthalate ester counterparts used as poly(vinyl chloride) (PVC) plasticizers. The bio derived plasticizers are comparable to the petroleum-derived incumbents in reducing the glass transition temperature and increasing the thermal stability of PVC. Furthermore, the dimethoxyterephthalic esters are expected to have an extended lifetime in the polymer matrix, due to their lower volatility and by lower diffusion coefficients calculated by molecular dynamic simulations. These results demonstrate that the isophthalate and phthalate components in polyesters and plasticizers, respectively, could be substituted with bio-based methoxyterephthalate derivatives.

Under nutrient starvation conditions, many microorganisms such as Pseudomonas putida store excess carbon in medium-chain-length polyhydroxyalkanoate ( mcl -PHA) granules. Biosynthesis of PHAs from the fatty acid biosynthetic pathway requires PhaG, which has long been thought to encode a 3-hydroxyacyl-ACP:CoA transferase, and PhaC, the PHA polymerase. Although this pathway has been extensively studied, the exact role of PhaG remains inconclusive. In this work we present in vitro biochemical and in vivo genetic evidence demonstrating that PhaG functions as a 3-hydroxyacyl-ACP thiolase, producing 3-hydroxyacids rather than 3-hydroxyacyl-CoA. We identified two CoA ligases, fadD1 and alkK , essential for conversion of mcl -3-hydroxyacids to 3-hydroxyacyl-CoA, and thus PHA production. Taken together, this redefines the PHA biosynthetic pathway to include PhaG-dependent hydrolysis of hydroxyacyl-ACP, ligation of the 3-hydroxyacid to coenzyme-A, and polymerization by PhaC. Using these insights, we engineered P. putida to produce 3.7 g/L extracellular 3-hydroxyacids, which can be used for chemical synthesis of performance-advantaged polymers. ### Competing Interest Statement AMG, WTW, JDH and DJP have filed a patent (U.S. Patent No. US20250154452A1) on mcl-3HA production application of the pathway described herein. United States Department of Energy, Agile BioFoundry, BOTTLE Consortium, Center for Nanophase Materials Sciences, Office of Science User Facility

Oxidative catalytic depolymerization of polystyrene (PS) can produce benzoic acid, but the annual consumption of benzoic acid is ~40 times lower than PS. For this catalytic oxidation method to be a viable means to manage PS waste, benzoic acid should be converted to higher-volume chemicals. We demonstrate a hybrid chemical and biological process that uses PS as feedstock for production of adipic acid, a high-volume co-monomer for nylon 6,6 via benzoic acid. Mn/Br co-catalyzed autoxidation of PS to benzoic acid proceeds with a yield of up to 94% in a solvent mixture of benzoic acid and water. The PS-derived benzoic acid undergoes bioconversion at near-quantitative yield to muconic acid, which is readily converted to adipic acid through catalytic hydrogenation. Process modeling, techno-economic analysis, and life cycle assessment estimate an adipic acid minimum selling price of $3.18/kg, with a 61% decrease in greenhouse gas emissions relative to production from fossil fuels.

ABSTRACT Vanillin is the most in‐demand flavouring compound in the world and because vanillin extracted from vanilla pods cannot meet the global demand, most vanillin on the market today is chemically synthesised. Increasing demands by consumers for natural ingredients have inspired efforts to develop vanillin derived from microbial sources. These efforts have been challenged by low titers, likely caused by the toxicity of vanillin to most microbial biocatalysts. In this study, we engineered a Pseudomonas putida KT2440‐derived strain that accumulated vanillin from ferulic acid to 0.64 g/L. To increase the overall titre, we applied a hydrophobic polystyrene‐based resin to vanillin‐accumulating cultures, which enabled an increase in total vanillin recovery to an apparent titre of 3.35 g/L. This study demonstrates that P. putida can accumulate vanillin from ferulic acid to higher titers when vanillin is removed from the cultivation medium, mitigating its toxicity.

An analysis method was developed for the quantitation of catabolic intermediates produced by white-rot fungi. Quantitation was performed using high pressure liquid chromatography paired with diode array detection (HPLC-DAD). Chromatographic separation was achieved using a mobile phase gradient to separate various aromatic analytes and muconic acid isomers on an HPLC reverse phase analytical column.

Feedstock variability represents a challenge in lignocellulosic biorefineries, as it can influence both lignocellulose deconstruction and microbial conversion processes for biofuels and biochemicals production. The impact of feedstock variability on microbial performance remains underexplored, and predictive tools for microbial behaviour are needed to mitigate risks in biorefinery scale-up. Here, twelve batches of corn stover were deconstructed via deacetylation, mechanical refining, and enzymatic hydrolysis to generate lignin-rich and sugar streams. These batches and their derived streams were characterised to identify their chemical components, and the streams were used as substrates for producing muconate and butyrate by engineered Pseudomonas putida and wildtype Clostridium tyrobutyricum, respectively. Bacterial performance (growth, product titers, yields, and productivities) differed among the batches, but no strong correlations were identified between feedstock composition and performance. To provide metabolic insights into the origin of these differences, we evaluated the effect of twenty-three isolated chemical components on these microbes, including three components in relevant bioprocess settings in bioreactors, and we found that growth-inhibitory concentrations were outside the ranges observed in the streams. Overall, this study generates a foundational dataset on P. putida and C. tyrobutyricum performance to enable future predictive models and underscores their resilience in effectively converting fluctuating lignocellulose-derived streams into bioproducts.

An analysis method was developed to allow for quantitation of muconic acid isomers (cis, cis and cis, trans) and aromatic compounds by ultra high pressure liquid chromatography paired with diode array detection (UHPLC-DAD). This was achieved by reproducible preparation of c,c and c,t isomers of muconic acid and chromatographic separation using a mobile phase gradient to separate various aromatic analytes and muconic isomers on a UHPLC reverse phase analytical column.

Efforts to produce aromatic monomers through catalytic lignin depolymerization have historically focused on aryl–ether bond cleavage. A large fraction of aromatic monomers in lignin, however, are linked by various carbon–carbon (C–C) bonds that are more challenging to cleave and limit the yields of aromatic monomers from lignin depolymerization. Here, we report a catalytic autoxidation method to cleave C–C bonds in lignin-derived dimers and oligomers from pine and poplar. The method uses manganese and zirconium salts as catalysts in acetic acid and produces aromatic carboxylic acids as primary products. The mixtures of the oxygenated monomers are efficiently converted to cis,cis -muconic acid in an engineered strain of Pseudomonas putida KT2440 that conducts aromatic O -demethylation reactions at the 4-position. This work demonstrates that autoxidation of lignin with Mn and Zr offers a catalytic strategy to increase the yield of valuable aromatic monomers from lignin.

An analytical method was developed using high performance liquid chromatography with refractive index detection (HPLC-RID) to quantify the concentration of sugars and carboxylic acids including formic acid, acetic acid, propionic acid, butyric acid, lactic acid, glucose, xylose, arabinose, and glycerol in aqueous samples. This list is non-exhaustive as this isocratic method with an acid modified mobile phase is compatible with analysis of many alcohols, small acids, and sugars. This method utilizes a Bio-Rad Aminex HPX-87H Ion Exclusion Column to provide chromatographic separation.

An analysis method was developed to quantify aromatic compounds present in lignin-rich, alkaline pre-treated liquors produced from corn stover. This method employs ultra-high pressure liquid chromatography paired with diode array detection (UHPLC-DAD). A reverse phase Luna Omega PS C18 column from Phenomenex and a gradient were used to separate various aromatic compounds.

An analytical method was developed using high performance liquid chromatography with refractive index detection (HPLC-RID) to quantify the concentration of ethylene glycol and 1,4-butanediol in aqueous samples. This list is non-exhaustive as this isocratic method with an acid modified mobile phase is compatible with analysis of many alcohols, small acids, and sugars. This method utilizes a Bio-Rad Aminex HPX-87H Ion Exclusion Column to provide chromatographic separation.

An analytical method was developed using high performance liquid chromatography with refractive index detection (HPLC-RID) to quantify the concentrations of aliphatic acids. The analytes quantified were oxalic acid, malic acid, malonic acid, succinic acid, glycolic acid, lactic acid, formic acid, acetic acid, and propionic acid in samples from lignin-derived streams. This method utilized an ion exclusion column to provide chromatographic separation across a 30 minute run-time.

Selective lignin depolymerization is a key step in lignin valorization to value-added products, and there are multiple catalytic methods to cleave labile aryl-ether bonds in lignin. However, the overall aromatic monomer yield is inherently limited by refractory carbon-carbon linkages, which are abundant in lignin and remain intact during most selective lignin deconstruction processes. In this work, we demonstrate that a Co/Mn/Br-based catalytic autoxidation method promotes carbon-carbon bond cleavage in acetylated lignin oligomers produced from reductive catalytic fractionation. The oxidation products include acetyl vanillic acid and acetyl vanillin, which are ideal substrates for bioconversion. Using an engineered strain of Pseudomonas putida, we demonstrate the conversion of these aromatic monomers to cis,cis-muconic acid. Overall, this study demonstrates that autoxidation enables higher yields of bioavailable aromatic monomers, exceeding the limits set by ether-bond cleavage alone.

Lignin valorization is being intensely pursued via tandem catalytic depolymerization and biological funneling to produce single products. In many lignin depolymerization processes, aromatic dimers and oligomers linked by carbon-carbon bonds remain intact, necessitating the development of enzymes capable of cleaving these compounds to monomers. Recently, the catabolism of erythro-1,2-diguaiacylpropane-1,3-diol (erythro-DGPD), a ring-opened lignin-derived β-1 dimer, was reported in Novosphingobium aromaticivorans. The first enzyme in this pathway, LdpA (formerly LsdE), is a member of the nuclear transport factor 2 (NTF-2)-like structural superfamily that converts erythro-DGPD to lignostilbene through a heretofore unknown mechanism. In this study, we performed biochemical, structural, and mechanistic characterization of the N. aromaticivorans LdpA and another homolog identified in Sphingobium sp. SYK-6, for which activity was confirmed in vivo. For both enzymes, we first demonstrated that formaldehyde is the C1 reaction product, and we further demonstrated that both enantiomers of erythro-DGPD were transformed simultaneously, suggesting that LdpA, while diastereomerically specific, lacks enantioselectivity. We also show that LdpA is subject to a severe competitive product inhibition by lignostilbene. Three-dimensional structures of LdpA were determined using X-ray crystallography, including substrate-bound complexes, revealing several residues that were shown to be catalytically essential. We used density functional theory to validate a proposed mechanism that proceeds via dehydroxylation and formation of a quinone methide intermediate that serves as an electron sink for the ensuing deformylation. Overall, this study expands the range of chemistry catalyzed by the NTF-2-like protein family to a prevalent lignin dimer through a cofactorless deformylation reaction.

An analysis method was developed to allow for quantitation of aromatic compounds by ultra high pressure liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) detection. This was achieved by using reverse phase chromatography and multiple reaction monitoring (MRM) in negative ion mode using electrospray ionization (ESI).

A downstream process was developed for producing pure monopotassium glucarate and crystalline glucaric acid from a fermentation broth.