Ultrahigh proton conductivity of four ionic hydrogen-bonded organic frameworks based on functionalized terephthalates.

Recently, the utilization of hydrogen-bonded organic frameworks (HOFs) with high crystallinity and inherent well-defined H-bonding networks in the field of proton conduction has received increasing attention, but obtaining HOFs with excellent water stability and prominent proton conductivity (σ) remains challenging. Herein, by employing functionalized terephthalic acids, 2,5-dihydroxyterephthalic acid, 2-hydroxyterephthalic acid, 2-nitro terephthalic acid, and terephthalic acid, respectively, four highly water-stable ionic HOFs (iHOFs), [(C8H5O6)(Me2NH2)]∙2H2O (iHOF 1), [(C8H5O5)(Me2NH2)] (iHOF 2), [(C8H4NO6)(Me2NH2)] (iHOF 3) and [(C8H5O4)(Me2NH2)] (iHOF 4) were efficiently prepared by a straightforward synthesis approach in DMF and H2O solutions. The alternating-current (AC) impedance testing in humid conditions revealed that all four iHOFs were temperature- and humidity-dependent σ, with the greatest value reaching 10-2 S·cm-1. As expected, the high density of free carboxylic acid groups, crystallization water, and protonated [Me2NH2]+ units offer adequate protons and hydrophilic environments for effective proton transport. Furthermore, the σ values of these iHOFs with different functional groups were compared. It was discovered that it dropped in the following order under 100 °C and 98 % relative humidity (RH): σ iHOF 1 (1.72 × 10-2 S·cm-1) > σ iHOF 2 (4.03 × 10-3 S·cm-1) > σ iHOF 3 (1.46 × 10-3 S·cm-1) > σ iHOF 4 (4.86 × 10-4 S·cm-1). Finally, we investigated the causes of the above differences and the proton transport mechanism inside the framework using crystal structure data, water contact angle tests, and activation energy values. This study provides new motivation to develop highly proton-conductive materials.

PBAT is biodegradable but what about the toxicity of its biodegradation products?

CONTEXT: Poly(butylene adipate-co-terephthalate) (PBAT) is a biodegradable plastic. It was introduced to the plastics market in 1998 and since then has been widely used around the world. The main idea of this research is to perform quantum chemical calculations to study the potential toxicity of PBAT and its degradation products. We analyzed the electron transfer capacity to determine its potential toxicity. We found that biodegradable products formed with benzene rings are as good electron acceptors as PBAT and OOH•. Our results indicate that the biodegradation products are potentially as toxic as PBAT. This might explain why biodegradation products alter the photosynthetic system of plants and inhibit their growth. From this and other previous investigations, we can think that biodegradable plastics could represent a potential environmental risk. METHODS: All DFT computations were performed using the Gaussian16 at M062x/6-311 + g(2d,p) level of theory without symmetry constraints. Electro-donating (ω-) and electro-accepting (ω +) powers were used as response functions.

A study on the performance, structure, composition, and release behavior changes of polybutylene adipate terephthalic acid (PBAT) film during food contact.

Polybutylene adipate terephthalic acid (PBAT) is an emerging biodegradable material in food packaging. However, concerns have been raised regarding the potential hazards it could pose to food safety. In this study, the changes of PBAT films during food contact and the release of small molecules were inestigated by a multiscale approach. On a macro-scale, the surface roughness of the films increased with the reduction in the concentration of food simulants and the increase in contact temperatures, especially after immersion in acidic food environments. On a micro-scale, the crystallinity (Xc) and degradation indexes (DI) of the films increased by 5.7-61.2% and 7.8-48.6%, respectively, which led to a decrease in thermal stability. On a scale approaching the molecular level, 2,4-di-tert-butylphenol (2,4-DTBP) was detected by gas chromatography-mass spectrometry (GC-MS/MS) with the highest migration content, and the release behavior of 2,4-DTBP was further investigated by migration kinetics. In addition, terephthalic acid (TPA), a hydrolysis product of PBAT, was detected in acidic food environments by liquid chromatography-mass spectrometry (LC-MS/MS). The results of this study could provide practical guidance and assistance to promote sustainable development in the field of food packaging.

Development of an internal two-stage upflow anaerobic reactor integrating biostimulation strategies to enhance the degradation of aromatic compounds in wastewater from purified terephthalic acid and dimethyl terephthalate manufacturing processes.

In this study, we aimed to establish high-rate biological treatment of purified terephthalic acid (PTA) and dimethyl terephthalate (DMT) wastewater that minimizes the inhibitory effects of high concentration benzoate and acetate. To achieve this, we developed a novel bioreactor system and biostimulation strategy. An internal two-stage upflow anaerobic (ITUA) reactor was operated with (i) a packed bed containing green tuff medium underlying (ii) a compartment seeded with anaerobic granular sludge. Ethylene glycol was amended to stimulate syntrophic interactions. Continuous operation of the system for 1,026 days achieve an organic removal rate of 11.0 ± 0.6 kg COD/m3/d. The abundance of aromatic degraders significantly increased during operation. Thus, we successfully developed a high-rate treatment system to treat wastewater from the PTA/DMT manufacturing processes by activating syntrophs in an ITUA reactor.

New combined absorption/1H NMR method for qualitative and quantitative analysis of PET degradation products.

Poly(ethylene terephthalate) (PET) is a very valuable and beneficial material for industrial purposes, with various different applications. Due to the high annual production volume of over 50 million tons worldwide and the indiscriminate disposal by consumers, the polymers accumulate in the environment, causing negative effects on various ecosystems. Biodegradation via suitable enzymes represents a promising approach to combat the plastic waste issue so validated methods are required to measure the efficiency and efficacy of these enzymes. PETase and MHETase from Ideonella sakaiensis are suitable enzymes needed in combination to completely degrade PET into its environmentally friendly monomers. In this project, we compare and combine a previously described bulk absorbance measurement method with a newly established 1H NMR analysis method of the PET degradation products mono(2-hydroxyethyl) terephthalic acid, bis(2-hydroxyethyl) terephthalic acid and terephthalic acid. Both were optimized regarding different solvents, pH values and drying processes. The accuracy of the measurements can be confirmed with sensitivity limits of 2.5-5 µM for the absorption method and 5-10 µM for the 1H NMR analysis. The combination of the described methods therefore allows a quantitative analysis by using bulk absorption coupled with a qualitative analysis through 1H NMR. The methods established in our work can potentially contribute to the development of suitable recycling strategies of PET using recombinant enzymes.

Luminescent lanthanide mixed N-phthaloylglycinate and terephthalate coordination polymers: Structural and optical properties.

A new class of lanthanide mixed-carboxylate ligands compounds with formula {[Ln2 (phthgly)4 (bdc)(H2 O)6 ]·(H2 O)4 }∞ , labelled as Ln3+ : Eu (1) and Gd (2) coordination polymers (CP) were synthesized under mild reaction conditions between lanthanide nitrate salts and a solution of N-phthaloylglycine (phthgly) and terephthalic (bdc) ligands. The (1) and (2) coordination polymers were formed by symmetric binuclear units, in which phthgly and bdc carboxylate ligands are coordinated to the lanthanide ions by different coordination modes. Surprisingly, all organic ligands participate in hydrogen bonding interactions, forming an extremally rigid crystalline structure. The red narrow emission bands from the 5 D0 →7 FJ transitions of the Eu3+ ion show a high colour purity. The intramolecular energy transfer process from L→Eu3+ ion has been discussed. The experimental intensity parameters (Ω2,4 ) reflect lower angular distortion and polarizability of the chemical environment around the metal ion compared with other Eu3+ compounds reported in the literature. This novel class of coordination polymer offers a more attractive platform for developing luminescent functional materials for different applications.

Biodegradation of terephthalic acid using Rhodococcus erythropolis MTCC 3951: Insights into the degradation process, applications in wastewater treatment and polyhydroxyalkanoate production.

Terephthalic acid (TPA) is an endocrine disruptor widely used as a plasticizer and as a monomer in the manufacturing of PET bottles. However, because of various harmful effects on humans and the environment, it is now recognized as a priority pollutant whose environmental level needs to be controlled. In the present work, the TPA biodegradation efficacy of the bacterium Rhodococcus erythropolis (MTCC 3951) was studied in mineral salt media with TPA as the sole carbon and energy source. R. erythropolis was observed to degrade 5 mM and 120 mM TPA within 10 h and 84 h of incubation, respectively. The degradation efficiency was further optimized by varying the culture conditions, and the following optimum conditions were obtained: inoculum size- 5% (v/v), temperature- 30 °C, agitation speed- 200 rpm, and pH- 8.0. The bacterium was found to use an ortho-cleavage pathway for TPA degradation determined based on enzymatic and GC-MS studies. Moreover, during the degradation of TPA, the bacterium was observed to produce polyhydroxyalkanoate (PHA)-a biopolymer. Biodegradation of 120 mM TPA resulted in an accumulation of PHA. The PHA granules were visualized using fluorescence and transmission electron microscopy and were later characterized using FTIR spectroscopy. Furthermore, the robustness of the bacterium was demonstrated by its ability to degrade TPA in real industrial wastewater. Overall, R. erythropolis (MTCC 3951) hold the potential for controlling TPA pollution in the environment and vis-à-vis the production of PHA biopolymer.

Engineering Comamonas testosteroni for the production of 2-pyrone-4,6-dicarboxylic acid as a promising building block.

BACKGROUND: Plastics are an indispensable part of our daily life. However, mismanagement at their end-of-life results in severe environmental consequences. The microbial conversion of these polymers into new value-added products offers a promising alternative. In this study, we engineered the soil-bacterium Comamonas testosteroni KF-1, a natural degrader of terephthalic acid, for the conversion of the latter to the high-value product 2-pyrone-4,6-dicarboxylic acid. RESULTS: In order to convert terephthalic acid to 2-pyrone-4,6-dicarboxylic acid, we deleted the native PDC hydrolase and observed only a limited amount of product formation. To test whether this was the result of an inhibition of terephthalic acid uptake by the carbon source for growth (i.e. glycolic acid), the consumption of both carbon sources was monitored in the wild-type strain. Both carbon sources were consumed at the same time, indicating that catabolite repression was not the case. Next, we investigated if the activity of pathway enzymes remained the same in the wild-type and mutant strain. Here again, no statistical differences could be observed. Finally, we hypothesized that the presence of a pmdK variant in the degradation operon could be responsible for the observed phenotype and created a double deletion mutant strain. This newly created strain accumulated PDC to a larger extent and again consumed both carbon sources. The double deletion strain was then used in a bioreactor experiment, leading to the accumulation of 6.5 g/L of product in 24 h with an overall productivity of 0.27 g/L/h. CONCLUSIONS: This study shows the production of the chemical building block 2-pyrone-4,6-dicarboxylic acid from terephthalic acid through an engineered C. testosteroni KF-1 strain. It was observed that both a deletion of the native PDC hydrolase as well as a pmdK variant is needed to achieve high conversion yields. A product titer of 6.5 g/L in 24 h with an overall productivity of 0.27 g/L/h was achieved.

Terephthalic Acid Intercalated CoNi-LDH Materials for Improved Li-O2 Battery.

CoNi-LDH (layered CoNi double hydroxides) hollow nanocages with specific morphology are obtained by Ni ion etching of ZIF-67 (Zeolitic imidazolate framework-67). The structure of the layered materials is further modified by molecular intercalation. The original interlayer anions are replaced by the ion exchange effect of terephthalic acid, which helps to increase the interlayer distance of the material. The intercalated cage-like structures not only benefit for the storage of oxygen, and the discharge product reaction, but also have more support between the material layers. The experimental results show that the excessive use of intercalation agent will affect structural stability of the intercalated CoNi-LDH. By adjusting the amount of terephthalic acid, the intercalated CoNi-LDH-2 (with 0.02 mmol terephthalic acid intercalated) is not easy to collapse after 209 cycles and shows the best electrochemical performance in Li-O2 battery.

Enzyme selection, optimization, and production toward biodegradation of post-consumer poly(ethylene terephthalate) at scale.

Poly(ethylene terephthalate) (PET) is one of the world's most widely used polyester plastics. Due to its chemical stability, PET is extremely difficult to hydrolyze in a natural environment. Recent discoveries in new polyester hydrolases and breakthroughs in enzyme engineering strategies have inspired enormous research on biorecycling of PET. This study summarizes our research efforts toward large-scale, efficient, and economical biodegradation of post-consumer waste PET, including PET hydrolase selection and optimization, high-yield enzyme production, and high-capacity enzymatic degradation of post-consumer waste PET. First, genes encoding PETase and MHETase from Ideonella sakaiensis and the ICCG variant of leaf-branch compost cutinase (LCCICCG ) were codon-optimized and expressed in Escherichia coli BL21(DE3) for high-yield production. To further lower the enzyme production cost, a pelB leader sequence was fused to LCCICCG so that the enzyme can be secreted into the medium to facilitate recovery. To help bind the enzyme on the hydrophobic surface of PET, a substrate-binding module in a polyhydroxyalkanoate depolymerase from Alcaligenes faecalis (PBM) was fused to the C-terminus of LCCICCG . The resulting four different LCCICCG variants (LCC, PelB-LCC, LCC-PBM, and PelB-LCC-PBM), together with PETase and MHETase, were compared for PET degradation efficiency. A fed-batch fermentation process was developed to produce the target enzymes up to 1.2 g L-1 . Finally, the best enzyme, PelB-LCC, was selected and used for the efficient degradation of 200 g L-1 recycled PET in a well-controlled, stirred-tank reactor. The results will help develop an economical and scalable biorecycling process toward a circular PET economy.

Towards Symmetric Thioamides: Microwave-Aided Synthesis of Terephthalic Acid Derivatives.

The multistep synthesis of novel bis-terephthalthioamides based on methyl esters of amino acids (AAs) was proposed using conventional heating and microwave-assisted approaches. In fact, the comparative case study on the thionation of new symmetrical diamides with Lawesson's reagent (LR) was performed. The microwave-accelerated small-scale methodology was successfully employed on the whole pathway from substrates (Gly, Ala, Val, Tyr, Ser) to products (symmetrical dithioamides of terephthalic acid), resulting in significantly reduced reaction time, energy requirements, and slightly increased reaction yields when compared to conventional heating. Moreover, the intermolecular similarity of novel terephthalic acid derivatives was estimated in the multidimensional space (mDS) of the structure/property-related in silico descriptors using principal component analysis (PCA) and hierarchical clustering analysis (HCA). The distance-oriented structure/property distribution was also correlated with the experimental lipophilic data.

Enzymatic post-consumer poly(ethylene terephthalate) (PET) depolymerization using commercial enzymes.

Poly(ethylene terephthalate) (PET) is a synthetic polymer widely used globally. The high PET resistance to biotic degradation and its improper destination result in the accumulation of this plastic in the environment, largely affecting terrestrial and aquatic animals. This work investigated post-consumer PET (PC-PET) degradation using five commercial hydrolase enzymes (Novozym 51032, CalB, Palatase, Eversa, Lipozyme TL). Humicola insolens cutinase (HiC, Novozym 51032) was the most active among the enzymes studied. Several important reaction parameters (enzyme type, dual enzyme system, enzyme concentration, temperature, ultrasound treatment) were evaluated in PC-PET hydrolysis using HiC. The concentration and the proportion (molar ratio) of hydrolysis products, terephthalic acid (TPA), mono(2-hydroxyethyl) terephthalate (MHET), and bis(2-hydroxyethyl) terephthalate (BHET), were significantly changed depending on the reaction temperature. The TPA released at 70 °C was 3.65-fold higher than at 50 °C. At higher temperatures, the conversion of MHET into TPA was favored. The enzymatic PET hydrolysis by HiC was very sensitive to the enzyme concentration, indicating that it strongly adsorbs on the polymer surface. The concentration of TPA, MHET, and BHET increased as the enzyme concentration increased, and a maximum was achieved using 40-50 vol % of HiC. The presented results add relevant data to optimizing enzyme-based PET recycling technologies.

Degradation characteristics of polybutylene adipate terephthalic acid (PBAT) and its effect on soil physicochemical properties: A comparative study with several polyethylene (PE) mulch films.

The degradation process of different types of mulch in agriculture and its effect on soil ecosystem should be considered comprehensively. To this end, the changes in performance, structure, morphology, and composition of PBAT film during the degradation process were examined through a multiscale approach in comparison with several PE films and their effects on the soil physicochemical properties were investigated. At the macroscopic scale, the load and elongation of all films decreased with increasing ages and depths. At the microscopic scale, the stretching vibration peak intensity (SVPI) for PBAT and PE films decreased by 48.8 âˆ¼ 60.2% and 9.3 âˆ¼ 38.6%, respectively. The crystallinity index (CI) increased by 67.3 âˆ¼ 209.6% and 15.6 âˆ¼ 21.8%, respectively. At the molecules scale, terephthalic acid (TPA) was detected in localized soil with PBAT mulch after 180 d. In short, the degradation characteristics of PE films were depended on their thickness and density. The PBAT film exhibited the highest degree of degradation. Simultaneously, the soil physicochemical properties such as soil aggregates, microbial biomass and pH were affected by the changes of film structure and components during the degradation process. This work has practical implications for the sustainable development of agriculture.

Sensitive detection of butyrylcholinesterase activity based on a stimuli-responsive fluorescence reaction.

A fluorogenic reaction between the chelate of Mn(II)-citric acid and terephthalic acid (PTA) was discovered, which was carried out through heating the aqueous mixture of Mn2+, citric acid and PTA. Detailed investigations indicated the reaction products were 2-hydroxyterephthalic acid (PTA-OH), which was attributed to the reaction between PTA and OH, formed by the triggering of Mn(II)-citric acid in the presence of dissolved O2. PTA-OH showed a strong blue fluorescence, peaked at 420 nm, and the fluorescence intensity presented a sensitive response to pH of the reaction system. Based on these mechanisms, the fluorogenic reaction was used for the detection of butyrylcholinesterase activity, achieving a detection limit of 0.15 U/L. The detection strategy was successfully applied in human serum samples, and it was also extended for the detection of organophosphorus pesticides and radical scavengers. Such a facile fluorogenic reaction and its stimuli-responsive properties offered an effective tool for designing detection pathways in the fields of clinical diagnosis, environmental monitoring and bioimaging.

Testicular toxicity in mice exposed to terephthalic acid in utero and during lactation.

Terephthalic acid (TPA) is a worldwide aromatic compound widely used to manufacture resins and the raw material for the polymerization reaction with ethylene glycol to produce polyethylene terephthalate, known as PET. The use of TPA extends to the synthesis of phthalates, plasticizers used in various industrialized products such as toys and cosmetics. The present study aimed to evaluate the testicular toxicity of terephthalic acid on male mice exposed in utero and during lactation to TPA in different developmental windows. The animals were treated intragastric with TPA at stock dispersal dosages corresponding to 0.0014 g/ml and 0.56 g/ml of TPA in 0.5% v/v carboxymethylcellulose as well as the control dose, composed solely of dispersion of carboxymethylcellulose (0.5% v/v). Four experimental windows were established: group I-treatment in utero, in the fetal period (gestational day-GD 10.5-18.5), with euthanasia at GD 18.5; group II-treatment in utero, in the fetal period (GD 10.5-18.5) and the lactational period (postnatal day (PND-15)), with euthanasia at 15 days; group III-treatment in utero in the fetal period (DG 10.5-18.5) with euthanasia at 70 days (age of sexual maturity, PND 70); group IV-treatment in utero, in the fetal period (GD 10.5-18.5) and the lactational period (PND-15), with euthanasia at 70 days (PND70). The results indicate that TPA changes the reproductive parameters (testicular weight, GI, penis size, and anogenital index) only at the dose of 0.56 g/ml in the fetal period. Data on the volumetric ratio of the testis elements show that the dispersion with the highest concentration of TPA significantly altered the blood vessel/capillary, lymphatic vessel, and connective tissue percentages. Only at the dose of 0.56 g/ml TPA was it effective in decreasing the Leydig and Sertoli cell numbers of the euthanized animals at GD 18.5. In group II, TPA increased the diameter and lumen of seminiferous tubules, which indicates that TPA accelerated the maturation process of Sertoli cells without changing the number and the nuclear volume of these cells. The Sertoli and Leydig cell numbers of the 70-day animals exposed to TPA in the gestational and lactational period were similar to the control. Therefore, the present study is the first in the literature to show that TPA presents a testicular toxicity during fetal (DG18.5) and postnatal life (PND15), without repercussion in adulthood (70 days).

Fungal Screening for Potential PET Depolymerization.

Approximately 400 billion PET bottles are produced annually in the world, of which from 8 to 9 million tons are discarded in oceans. This requires developing strategies to urgently recycle them. PET recycling can be carried out using the microbial hydrolysis of polymers when monomers and oligomers are released. Exploring the metabolic activity of fungi is an environmentally friendly way to treat harmful polymeric waste and obtain the production of monomers. The present study addressed: (i) the investigation of potential of strains with the potential for the depolymerization of PET bottles from different manufacturers (crystallinity of 35.5 and 10.4%); (ii) the search for a culture medium that favors the depolymerization process; and (iii) gaining more knowledge on fungal enzymes that can be applied to PET recycling. Four strains (from 100 fungal strains) were found as promising for conversion into terephthalic acid from PET nanoparticles (npPET): Curvularia trifolii CBMAI 2111, Trichoderma sp. CBMAI 2071, Trichoderma atroviride CBMAI 2073, and Cladosporium cladosporioides CBMAI 2075. The fermentation assays in the presence of PET led to the release of terephthalic acid in concentrations above 12 ppm. Biodegradation was also confirmed using mass variation analyses (reducing mass), scanning electron microscopy (SEM) that showed evidence of material roughness, FTIR analysis that showed band modification, enzymatic activities detected for lipase, and esterase and cutinase, confirmed by monomers/oligomers quantification using high performance liquid chromatography (HPLC-UV). Based on the microbial strains PET depolymerization, the results are promising for the exploration of the selected microbial strain.

Call for biotechnological approach to degrade plastic in the era of COVID-19 pandemic.

Plastic pollution is a global issue and has become a major concern since Coronavirus disease (COVID)-19. In developing nations, landfilling and illegal waste disposal are typical ways to dispose of COVID-19-infected material. These technologies worsen plastic pollution and other human and animal health problems. Plastic degrades in light and heat, generating hazardous primary and secondary micro-plastic. Certain bacteria can degrade artificial polymers using genes, enzymes, and metabolic pathways. Microorganisms including bacteria degrade petrochemical plastics slowly. High molecular weight, strong chemical bonds, and excessive hydrophobicity reduce plastic biodegradation. There is not enough study on genes, enzymes, and bacteria-plastic interactions. Synthetic biology, metabolic engineering, and bioinformatics methods have been created to biodegrade synthetic polymers. This review will focus on how microorganisms' degrading capacity can be increased using recent biotechnological techniques.

p-Xylene Oxidation to Terephthalic Acid: New Trends.

Large-scale terephthalic acid production from the oxidation of p-xylene is an especially important process in the polyester industry, as it is mainly used in polyethylene terephthalate (PET) manufacturing, a polymer that is widely used in fibers, films, and plastic products. This review presents and discusses catalytic advances and new trends in terephthalic acid production (since 2014), innovations in terephthalic acid purification processes, and simulations of reactors and reaction mechanisms.

Cd2+ and Zn2+ Complexes with 2,4,6-Tri(2-pyridyl)-s-Triazine and Terephthalate for Rapid, High-Capacity Adsorption of Congo Red.

Three crystal complexes were designed and synthesised through the solvothermal method, with Cu2+ , Zn2+ , and Cd2+ ions as the metal centres and 2,4,6-tri(2-pyridyl)-s-triazine (TPTZ) and terephthalate (BDC2- ) as the ligands. Their compositions were determined to be Cd(TPTZ)Cl2 (Cd-MOF), {[Zn(TPTZ)(BDC)] ⋅ 3H2 O}n (Zn-MOF), and Cu2 (PCA)2 (BDC)(H2 O)2 (Cu-MOF) (PCA- =2-pyridinium amide), respectively. Cd-MOF can adsorb 90 % of Congo red (CR) in 10 s at room temperature and atmospheric pressure, and CR removal was complete at 20 s over a wide pH range. The adsorption capacity for CR reached 1440 mg g-1 in 5 min. Selective adsorption was demonstrated in mixed dyes. The adsorption kinetic data agree well with the pseudo-second-order kinetic model. The Temkin model was successfully used to evaluate the adsorption isotherms of CR on Cd-MOF at room temperature, suggesting that adsorption occurs through a hybrid of monolayer and multilayer mechanisms.

Improved terephthalic acid production from p-xylene using metabolically engineered Pseudomonas putida.

Terephthalic acid (TPA) is an important commodity chemical used as a monomer of polyethylene terephthalate (PET). Since a large quantity of PET is routinely manufactured and consumed worldwide, the development of sustainable biomanufacturing processes for its monomers (i.e. TPA and ethylene glycol) has recently gained much attention. In a previous study, we reported the development of a metabolically engineered Escherichia coli strain producing 6.7 g/L of TPA from p-xylene (pX) with a productivity and molar conversion yield of 0.278 g/L/h and 96.7 mol%, respectively. Here, we report metabolic engineering of Pseudomonas putida KT2440, a microbial chassis particularly suitable for the synthesis of aromatic compounds, for improved biocatalytic conversion of pX to TPA. To develop a plasmid-free, antibiotic-free, and inducer-free biocatalytic process for cost-competitive TPA production, all heterologous genes required for the synthetic pX-to-TPA bioconversion pathway were integrated into the chromosome of P. putida KT2440 by RecET-based markerless recombineering and overexpressed under the control of constitutive promoters. Next, TPA production was enhanced by integrating multiple copies of the heterologous genes to the ribosomal RNA genes through iteration of recombineering-based random integration and subsequent screening of high-performance strains. Finally, fed-batch fermentation process was optimized to further improve the performance of the engineered P. putida strain. As a result, 38.25 ± 0.11 g/L of TPA was produced from pX with a molar conversion yield of 99.6 ± 0.6%, which is equivalent to conversion of 99.3 ± 0.8 g pX to 154.6 ± 0.5 g TPA. This superior pX-to-TPA biotransformation process based on the engineered P. putida strain will pave the way to the commercial biomanufacturing of TPA in an industrial scale.