Sorbate metal complexes as newer antibacterial, antibiofilm, and anticancer compounds.

BACKGROUND: The ineffectiveness of treatments for infections caused by biofilm-producing pathogens and human carcinoma presents considerable challenges for global public health organizations. To tackle this issue, our study focused on exploring the potential of synthesizing new complexes of Co(II), Cu(II), Ni(II), and Zn(II) with sorbic acid to enhance its antibacterial, antibiofilm, and anticancer properties. METHODS: Four novel complexes were synthesized as solid phases by reacting sorbic acid with Co(II), Cu(II), Ni(II), and Zn(II). These complexes were characterized by various technique, including infrared spectra, UV-Visible spectroscopy, proton nuclear magnetic resonance (1H NMR), and thermal analysis techniques, including thermogravimetry (TG). RESULTS: The data acquired from all investigated chemical characterization methods confirmed the chemical structure of the sorbate metal complexes. These complexes exhibited antibacterial and antibiofilm properties against both Gram-positive and Gram-negative bacteria. Furthermore, these complexes enhanced the antibacterial effects of commonly used antibiotics, such as gentamicin and imipenem, with fractional inhibitory concentration (FIC) indices ≤ 0.5. Notably, the Cu(II) complex displayed the most potent antibacterial and antibiofilm activities, with minimum inhibitory concentration (MIC) values of 312.5 µg/mL and 625.0 µg/mL for Bacillus cereus and Escherichia coli, respectively. Additionally, in vitro assays using the methyl thiazolyl tetrazolium (MTT) method showed inhibitory effects on the growth of the human colon carcinoma cell line (HCT-116 cells) following treatment with the investigated metal complexes. The IC50 values for Co(II), Cu(II), Zn(II), and Ni(II) were 3230 µg/mL, 2110 µg/mL, 3730 µg/mL, and 2240 µg/mL, respectively. CONCLUSION: Our findings offer potential for pharmaceutical companies to explore the development of novel combinations involving traditional antibiotics or anticancer drugs with sorbate copper complex.

Biosensors for the detection of chorismate and cis,cis-muconic acid in Corynebacterium glutamicum.

Corynebacterium glutamicum ATCC 13032 is a promising microbial chassis for industrial production of valuable compounds, including aromatic amino acids derived from the shikimate pathway. In this work, we developed two whole-cell, transcription factor based fluorescent biosensors to track cis,cis-muconic acid (ccMA) and chorismate in C. glutamicum. Chorismate is a key intermediate in the shikimate pathway from which value-added chemicals can be produced, and a shunt from the shikimate pathway can divert carbon to ccMA, a high value chemical. We transferred a ccMA-inducible transcription factor, CatM, from Acinetobacter baylyi ADP1 into C. glutamicum and screened a promoter library to isolate variants with high sensitivity and dynamic range to ccMA by providing benzoate, which is converted to ccMA intracellularly. The biosensor also detected exogenously supplied ccMA, suggesting the presence of a putative ccMA transporter in C. glutamicum, though the external ccMA concentration threshold to elicit a response was 100-fold higher than the concentration of benzoate required to do so through intracellular ccMA production. We then developed a chorismate biosensor, in which a chorismate inducible promoter regulated by natively expressed QsuR was optimized to exhibit a dose-dependent response to exogenously supplemented quinate (a chorismate precursor). A chorismate-pyruvate lyase encoding gene, ubiC, was introduced into C. glutamicum to lower the intracellular chorismate pool, which resulted in loss of dose dependence to quinate. Further, a knockout strain that blocked the conversion of quinate to chorismate also resulted in absence of dose dependence to quinate, validating that the chorismate biosensor is specific to intracellular chorismate pool. The ccMA and chorismate biosensors were dually inserted into C. glutamicum to simultaneously detect intracellularly produced chorismate and ccMA. Biosensors, such as those developed in this study, can be applied in C. glutamicum for multiplex sensing to expedite pathway design and optimization through metabolic engineering in this promising chassis organism. ONE-SENTENCE SUMMARY: High-throughput screening of promoter libraries in Corynebacterium glutamicum to establish transcription factor based biosensors for key metabolic intermediates in shikimate and ß-ketoadipate pathways.

Leukemia risk assessment of exposure to low-levels of benzene based on the linearized multistage model.

Objectives: To assess leukemia risk in occupational populations exposed to low levels of benzene. Methods: Leukemia incidence data from the Chinese Benzene Cohort Study were fitted using the Linearized multistage (LMS) model. Individual benzene exposure levels, urinary S-phenylmercapturic acid (S-PMA) and trans, trans-muconic acid (t, t-MA) were measured among 98 benzene-exposed workers from factories in China. Subjects were categorized into four groups by rounding the quartiles of cumulative benzene concentrations (< 3, 3-5, 5-12, ≥12 mg/m3·year, respectively). The risk of benzene-induced leukemia was assessed using the LMS model, and the results were validated using the EPA model and the Singapore semi-quantitative risk assessment model. Results: The leukemia risks showed a positive correlation with increasing cumulative concentration in the four exposure groups (excess leukemia risks were 4.34, 4.37, 4.44 and 5.52 × 10-4, respectively; Ptrend < 0.0001) indicated by the LMS model. We also found that the estimated leukemia risk using urinary t, t-MA in the LMS model was more similar to those estimated by airborne benzene compared to S-PMA. The leukemia risk estimated by the LMS model was consistent with both the Singapore semi-quantitative risk assessment model at all concentrations and the EPA model at high concentrations (5-12, ≥12 mg/m3·year), while exceeding the EPA model at low concentrations (< 3 and 3-5 mg/m3·year). However, in all four benzene-exposed groups, the leukemia risks estimated by these three models exceeded the lowest acceptable limit for carcinogenic risk set by the EPA at 1 × 10-6. Conclusion: This study demonstrates the utility of the LMS model derived from the Chinese benzene cohort in assessing leukemia risk associated with low-level benzene exposure, and suggests that leukemia risk may occur at cumulative concentrations below 3 mg/m3·year.

Association between urinary BTEX metabolites and dyslexic odds among school-aged children.

Benzene, toluene, ethylbenzene, and xylene (BTEX) are ubiquitous in the environment, and all of them can cause neurotoxicity. However, the association between BTEX exposure and dyslexia, a disorder with language network-related regions in left hemisphere affected, remains unclear. We aimed to assess the relationship between BTEX exposure and dyslexic odds among school-aged children. A case-control study, including 355 dyslexics and 390 controls from three cities in China, was conducted. Six BTEX metabolites were measured in their urine samples. Logistic regression model was used to explore the association between the BTEX metabolites and the dyslexic odds. Urinary trans,trans-muconic acid (MU: a metabolite of benzene) was significantly associated with an increased dyslexic odds [odds ratio (OR) = 1.23, 95% confidence interval (CI): 1.01, 1.50], and the adjusted OR of the dyslexic odds in the third tertile was 1.72 (95% CI: 1.06, 2.77) compared to that in the lowest tertile regarding urinary MU concentration. Furthermore, the association between urinary MU level and the dyslexic odds was more pronounced among children from low-income families based on stratified analyses. Urinary metabolite levels of toluene, ethylbenzene, and xylene were not found to be associated with the dyslexic odds. In summary, elevated MU concentrations may be associated with an increased dyslexic odds. We should take measures to reduce MU related exposure among children, particularly those with low family income.

Optimization of benzene exposure risk assessment: An integrated approach utilizing internal and external concentrations with a focus on biomarkers S-PMA & t, t-MA.

In the majority of occupational settings within China, the concentrations of benzene are observed to fall markedly below the demarcated detection thresholds. Employing traditional risk assessment models, the presence of exceptionally low airborne benzene exposure concentrations may infuse heightened degrees of uncertainty. Consequently, the necessity arises to investigate risk assessment methodologies more apt for the prevalent exposure environment among employees. In the present study, a pharmacokinetic model premised on urinary benzene metabolites (S-PMA and t, t-MA) was employed to ascertain a more precise daily airborne benzene exposure concentration per individual. This value was integrated into the linear multistage model as the 'internal exposure concentration'. In conjunction with the U.S National Environmental Protection Agency's (EPA) inhalation risk assessment model predicated on the external exposure concentration, the Singapore Ministry of Manpower's (MOM) model, and the linear multistage (LMS) model, the carcinogenic and non-carcinogenic effects of benzene were evaluated for 1781 benzene-exposed employees across 76 enterprises in Jiangsu Province. Findings suggest that in the linear multilevel model assessment, the cancer risk levels based on t, t-MA and S-PMA were higher in the printing and recording media reproduction industry, automobile manufacturing industry, general equipment manufacturing industry and the furniture manufacturing industry (median 2.842 × 10-4, 2.819 × 10-4, 2.809 × 10-4, and 2.678 × 10-4), which align more consistently with the actual benzene exposure circumstances of each industry's study participants, with overall risk levels calculated by the linear multistage model exceeding those of the EPA inhalation risk assessment model and the MOM model. This implies that the linear multistage model of internal exposure, based on the reciprocal of benzene biomarkers S-PMA and t, t-MA for airborne benzene exposure, presents enhanced sensitivity and suitability for the current occupational health risk assessment of workers. Without doubt, biomarker-based benzene exposure risk assessment emerges as the optimal choice.

Multi-functional polyvinyl alcohol/tannin acid composite films incorporated with lignin nanoparticles loaded by potassium sorbate.

The biocompatible, biodegradable and strong polyvinyl alcohol-based films have been widely investigated and used in the field of active packaging. To endow with diverse function, this paper firstly prepared lignin nanoparticles loaded with potassium sorbate (LNP@PS) as additives to exploit additional antibacterial, UV blocking, oxygen barrier, and water barrier properties. Besides, tannin acid (TA) was incorporated for compensating and further enhancing mechanical properties. Results showed that the PVA-based composite films containing 3 % LNP@PS and 5 % TA could achieve the optimal tensile strength at 74.51 MPa, water vapor permeability at 7.015·10-13·g·cm/cm2·s·Pa and oxygen permeability at 1.93 cm3/m2·24 h MPa, which was an 165 % of increase, 47 % and 112 % of reduction respectively compared to pure PVA films. Additionally, the composite films exhibited apparently superior bacteria and oxygen resistance properties evidenced by microbial infection and free radical scavenging performance. In addition, the slow-release effect of PS assisted the strawberry preservation with an extension of 3 days, which provided a promising novel route to prepare active food packaging material.

Genome-wide host-pathway interactions affecting cis-cis-muconic acid production in yeast.

The success of forward metabolic engineering depends on a thorough understanding of the behaviour of a heterologous metabolic pathway within its host. We have recently described CRI-SPA, a high-throughput gene editing method enabling the delivery of a metabolic pathway to all strains of the Saccharomyces cerevisiae knock-out library. CRI-SPA systematically quantifies the effect of each modified gene present in the library on product synthesis, providing a complete map of host:pathway interactions. In its first version, CRI-SPA relied on the colour of the product betaxanthins to quantify strains synthesis ability. However, only a few compounds produce a visible or fluorescent phenotype limiting the scope of our approach. Here, we adapt CRI-SPA to onboard a biosensor reporting the interactions between host genes and the synthesis of the colourless product cis-cis-muconic acid (CCM). We phenotype >9,000 genotypes, including both gene knock-out and overexpression, by quantifying the fluorescence of yeast colonies growing in high-density agar arrays. We identify novel metabolic targets belonging to a broad range of cellular functions and confirm their positive impact on CCM biosynthesis. In particular, our data suggests a new interplay between CCM biosynthesis and cytosolic redox through their common interaction with the oxidative pentose phosphate pathway. Our genome-wide exploration of host:pathway interaction opens novel strategies for improved production of CCM in yeast cell factories.

Corynebacterium glutamicum cell factory design for the efficient production of cis, cis-muconic acid.

Cis, cis-muconic acid (MA) is widely used as a key starting material in the synthesis of diverse polymers. The growing demand in these industries has led to an increased need for MA. Here, we constructed recombinant Corynebacterium glutamicum by systems metabolic engineering, which exhibit high efficiency in the production of MA. Firstly, the three major degradation pathways were disrupted in the MA production process. Subsequently, metabolic optimization strategies were predicted by computational design and the shikimate pathway was reconstructed, significantly enhancing its metabolic flux. Finally, through optimization and integration of key genes involved in MA production, the recombinant strain produced 88.2 g/L of MA with the yield of 0.30 mol/mol glucose in the 5 L bioreactor. This titer represents the highest reported titer achieved using glucose as the carbon source in current studies, and the yield is the highest reported for MA production from glucose in Corynebacterium glutamicum. Furthermore, to enable the utilization of more cost-effective glucose derived from corn straw hydrolysate, we subjected the strain to adaptive laboratory evolution in corn straw hydrolysate. Ultimately, we successfully achieved MA production in a high solid loading of corn straw hydrolysate (with the glucose concentration of 83.56 g/L), resulting in a titer of 19.9 g/L for MA, which is 4.1 times higher than that of the original strain. Additionally, the glucose yield was improved to 0.33 mol/mol. These provide possibilities for a greener and more sustainable production of MA.

Associations of personal urinary volatile organic compounds and lung function in children.

BACKGROUND: We investigated the correlation between urine VOC metabolites and airway function in children exposed to anthropogenic volatile organic compounds (VOCs), notable pollutants impacting respiratory health. METHODS: Out of 157 respondents, 141 completed skin prick tests, spirometry, IOS, and provided urine samples following the International Study of Asthma and Allergies in Childhood (ISAAC)-related questions. Allergic sensitization was assessed through skin prick tests, and airway functions were evaluated using spirometry and impulse oscillometry (IOS). Forced expiratory volume in 1 s (FEV1), forced vital capacity (FVC) was recorded and FEV1/FVC ratio was calculated. Airway mechanics parameters including respiratory resistance at 5 Hz (Rrs5) mean respiratory resistance between 5 Hz and 20 Hz (Rrs5-20), were also recorded. Urine concentrations of metabolites of benzene, ethylbenzene, toluene, xylene, styrene, formaldehyde, carbon-disulfide were analyzed by gas chromatography/tandem mass spectroscopy. RESULTS: The median age at study participation was 7.1 (SD 0.3) years. Muconic acid (benzene metabolites) and o-methyl-hippuric acid (xylene metabolites) above medians were associated with a significant increase in Rrs5 (muconic acid: aß = 0.150, p = .002; o-methyl-hippuric acid: aß = 0.143, p = .023) and a decrease in FEV1/FVC (o-methyl-hippuric acid: aß = 0.054, p = .028) compared to those below median. No associations were observed for Rrs5-20 and FEV1 between the groups categorized as above and below the median (all parameter p values > .05). CONCLUSIONS: Elevated levels of benzene and xylene metabolites were associated with a significant increase in Rrs5 and a decrease in FEV1/FVC, related to increased resistance and restrictive lung conditions compared to individuals with concentrations below the median.

Effects of potassium sorbate on systemic inflammation and gut microbiota in normal mice: A comparison of continuous intake and washout period.

Potassium sorbate (PS) is a widely used food preservative in the field of food industry. However, the effects of continuous intake and washout period of PS on host health are still unclear. In this study, to investigate long-term effect and after-effect of different concentrations and time points of PS, healthy mice were orally exposed to 150 mg/kg, 500 mg/kg and 1000 mg/kg of PS for 10 weeks, and washout treatment for another 5 weeks, respectively. The results indicated that PS intake for 10 weeks had no obvious effects on organs and adipose tissue, nor did it noteworthily interfere with glucolipid metabolism in the serum. However, it caused inflammatory cell infiltration in the liver, increased serum interleukin (IL)-1ß level, changed abundances of gut microbiota but failed to promote the production of short chain fatty acids in the gut. After washout period for 5 weeks, liver inflammation and IL-1ß level were decreased, and gut environment developed towards a healthier condition. Specifically, PS washout significantly increased abundance of Lachnospiraceae_NK4A136_group and the production of isobutyric acid. This study confirmed washout period eliminated negative effects from continuous intake of PS, which provided positive evidence for its safety.

Utilizing functional genomics in Saccharomyces cerevisiae to characterize food preservative compounds: A pilot study.

Chemical preservatives are ubiquitously used to suppress the growth of or kill microorganisms across numerous industries, including the food industry. Utilizing yeast functional genomic techniques, genes and their functions can be observed at a genomic scale to elucidate how environmental stressors (e.g., chemical preservatives) impact microbial survival. These types of chemical genomics approaches can reveal genetic mutations that result in preservative resistance or sensitivity, assist in identification of preservative mechanism of action, and can be used to compare different preservatives for rational design of preservative mixtures. In this proof-of-concept study, we performed deletion and high-copy genetic expression screens to identify mutants that confer drug resistance to sodium benzoate, potassium sorbate, rosemary extract, and Natamax. By observing overlapping mutant genes between genetic screens, we were able to identify functional overlap between chemical preservatives and begin to explain mechanisms of action for these compounds.

Molecular docking study of frequently used food additives for selected targets depending on the chromosomal abnormalities they cause.

Food additives (FAs) (flavor enhancers, sweeteners, etc.) protect foods during storage and transportation, making them attractive to consumers. Today, while the desire to access natural foods is increasing, the chemicals added to foods have started to be questioned. In this respect, genotoxicity tests have gained importance. Studies show that some food additives may have genotoxic risks. Previous studies carried out in our laboratory also revealed genotoxic effects of Monopotassium glutamate (MPG), Monosodium glutamate (MSG), Magnesium diglutamate (MDG) as flavor enhancers; Potassium benzoate (PB), Potassium sorbate (PS), Sodium benzoate (SB), Sodium sorbate (SS) as preservatives; Acesulfame potassium (ACE-K), Xylitol (XYL) as sweeteners. In this study, we determined the interactions of these food additives with ATM and p53 proteins, which are activated in the cell due to genotoxic effects, and with DNA by employing the molecular docking method for the first time. Among the food additives, SB (-4.307) for ATM, XYL (-4.629) for p53, and XYL (-4.927) for DNA showed the highest affinity. Therefore, flexible docking (IFD) scores were determined for SB, XYL, and MDG from flavor enhancers. The potential binding modes of the food additives to target molecules' possible inhibition mechanisms were determined by molecular docking. Thus, new information was obtained to show how these additives cause chromosomal abnormalities.

Engineering Novosphingobium aromaticivorans to produce cis,cis-muconic acid from biomass aromatics.

The platform chemical cis,cis-muconic acid (ccMA) provides facile access to a number of monomers used in the synthesis of commercial plastics. It is also a metabolic intermediate in the ß-ketoadipic acid pathway of many bacteria and, therefore, a current target for microbial production from abundant renewable resources via metabolic engineering. This study investigates Novosphingobium aromaticivorans DSM12444 as a chassis for the production of ccMA from biomass aromatics. The N. aromaticivorans genome predicts that it encodes a previously uncharacterized protocatechuic acid (PCA) decarboxylase and a catechol 1,2-dioxygenase, which would be necessary for the conversion of aromatic metabolic intermediates to ccMA. This study confirmed the activity of these two enzymes in vitro and compared their activity to ones that have been previously characterized and used in ccMA production. From these results, we generated one strain that is completely derived from native genes and a second that contains genes previously used in microbial engineering synthesis of this compound. Both of these strains exhibited stoichiometric production of ccMA from PCA and produced greater than 100% yield of ccMA from the aromatic monomers that were identified in liquor derived from alkaline pretreated biomass. Our results show that a strain completely derived from native genes and one containing homologs from other hosts are both capable of stoichiometric production of ccMA from biomass aromatics. Overall, this work combines previously unknown aspects of aromatic metabolism in N. aromaticivorans and the genetic tractability of this organism to generate strains that produce ccMA from deconstructed biomass.IMPORTANCEThe production of commodity chemicals from renewable resources is an important goal toward increasing the environmental and economic sustainability of industrial processes. The aromatics in plant biomass are an underutilized and abundant renewable resource for the production of valuable chemicals. However, due to the chemical composition of plant biomass, many deconstruction methods generate a heterogeneous mixture of aromatics, thus making it difficult to extract valuable chemicals using current methods. Therefore, recent efforts have focused on harnessing the pathways of microorganisms to convert a diverse set of aromatics into a single product. Novosphingobium aromaticivorans DSM12444 has the native ability to metabolize a wide range of aromatics and, thus, is a potential chassis for conversion of these abundant compounds to commodity chemicals. This study reports on new features of N. aromaticivorans that can be used to produce the commodity chemical cis,cis-muconic acid from renewable and abundant biomass aromatics.

Using labelling information to evaluate the distribution of food additives in products marketed in Brazil.

Food additives are ingredients added to food and beverages in order to modify physical, chemical, biological or sensory characteristics, contributing to the development of products that are safer and more convenient and attractive. Although they are widely used by food industries, little is known about the distribution of the food additives currently employed in products marketed in Brazil. Therefore, this study aimed to use labelling information of commercial food and beverages in order to identify the substances currently used by the Brazilian industry. For this, 3300 labels of 426 food items described in the Personal Food Consumption module of the Household Budget Survey (POF 2017/2018) of the Brazilian Institute of Geography and Statistics (IBGE) were searched on websites of supermarket chains and food companies as well as by visits to commercial establishments. According to the ingredient lists of the evaluated samples, 186 distinct food additives were identified. The average number of declared substances per product was between 4 and 5, with most of the labels indicating the presence of 2 or 3 additives. Considering the identified substances, 93 had an Acceptable Daily Intake (ADI) not specified or not limited and 89 were additives with a numerical ADI. Citric acid and lecithin were the most frequent substances with ADI not specified or not limited while potassium sorbate and pentasodium triphosphate were the most frequent food additives with numerical ADI. The most frequent combinations found in the evaluated samples were of potassium sorbate and citric acid, and also sodium nitrite, sodium erythorbate, and sodium diphosphate. This study demonstrates the distribution of food additives in products marketed in Brazil, allowing the creation of a comprehensive and unprecedented database, which will contribute to the evaluation of usage trends and future exposure studies for health risk assessment.

Benzoates and in situ formed benzene in food supplements and risk assessment.

Upward trend in the use of food supplements urged the assessment of their safety. Eighty-eight liquid herbal supplements collected in Novi Sad (Serbia) in 2018 (36 samples) and 2021 (52 samples) were analysed for the presence of benzoates and sorbates (HPLC-UV) and benzene (HS-GC/MS). Benzoic acid varied from 599 to 9253 mg/kg and sorbic acid between 185 and 1658 mg/kg. The acceptable daily intake of sorbic acid was not reached, but in case of benzoic acid, it was exceeded by 5.3% of the samples. The presence of benzene was confirmed in 41.2% of benzoate preserved supplements (0.9-51.7 µg/kg). Benzene exposure revealed no health concern: maximum hazard quotients ranged from 0.39% (toddlers) to 0.84% (adolescents); minimum margins of exposure were between 35,680 (adolescents) and 77,419 (toddlers); estimates of lifetime cancer risk did not reach one extra cancer case per 100 000 persons. However, measures to mitigate benzene presence in food should be considered.

Inactivation of Bacillus subtilis spores by a combination of high-pressure thermal treatment and potassium sorbate.

Combining High-pressure Thermal Treatment (HPTT) and Potassium Sorbate (PS) may have a stronger spore inactivation effect. Spores of Bacillus subtilis were subjected to HPTT at 600 MPa-65 °C/75 °C and a combination of HPTT and PS of 0.1% and 0.2% concentrations. After these treatments, different procedures and techniques were employed to investigate the spore's inactivation. The results revealed that 4.92 ± 0.05 log spores were inactivated after treatment at 600 MPa-75 °C, while 5.97 ± 0.09 log spores were inactivated when the HPTT treatment was combined with 0.2% PS. Changes in permeability of the spore's inner membrane were characterized by OD600 value and release rates of nucleic acids, protein, and dipicolinic acid (DPA). Compared with HPTT treatment at 600 MPa-75 °C, the OD600 value of spores decreased further by about 50% after treatment with a combination of HPTT and 0.2% PS. Additionally, the combined treatments resulted in a significant increase in the OD260 and OD280 values, as well as the DPA release. The spore size analysis indicated a significant decrease in the size of spores treated with a combination of HPTT at 600 MPa-75 °C and PS of 0.2% concentration. Furthermore, the flow cytometry analysis and confocal laser scanning microscopy (CLSM) analysis indicated that the inner membrane damage of spores was higher after combined treatments than that after HPTT treatment alone. A significant reduction was also found in the Na+/K+-ATPase activity after the combined treatments. Also, the FTIR analysis revealed that the combined treatments resulted in significant adverse changes in the spores' inner membrane, cell wall, cortex, and nucleic acid. Therefore, the combination of HPTT and PS has a stronger inactivation effect and can be suggested as a promising strategy for the inactivation of Bacillus subtilis spores.

Screening non-conventional yeasts for acid tolerance and engineering Pichia occidentalis for production of muconic acid.

Saccharomyces cerevisiae is a workhorse of industrial biotechnology owing to the organism's prominence in alcohol fermentation and the suite of sophisticated genetic tools available to manipulate its metabolism. However, S. cerevisiae is not suited to overproduce many bulk bioproducts, as toxicity constrains production at high titers. Here, we employ a high-throughput assay to screen 108 publicly accessible yeast strains for tolerance to 20 g L-1 adipic acid (AA), a nylon precursor. We identify 15 tolerant yeasts and select Pichia occidentalis for production of cis,cis-muconic acid (CCM), the precursor to AA. By developing a genome editing toolkit for P. occidentalis, we demonstrate fed-batch production of CCM with a maximum titer (38.8 g L-1), yield (0.134 g g-1 glucose) and productivity (0.511 g L-1 h-1) that surpasses all metrics achieved using S. cerevisiae. This work brings us closer to the industrial bioproduction of AA and underscores the importance of host selection in bioprocessing.

Evaluating the potential of natural product combinations with sorbic acid for improving preservative action against food-spoilage yeasts.

Fungal control methods commonly involve the use of antifungals or preservatives, which can raise concerns about broader effects of these stressors on non-target organisms, spread of resistance and regulatory hurdles. Consequently, control methods enabling lower usage of such stressors are highly sought, for example chemical combinations that synergistically inhibit target-organisms. Here, we investigated how well such a principle extends to improving efficacy of an existing but tightly controlled food preservative, sorbic acid. A screen of ∼200 natural products for synergistic fungal inhibition in combinations with sorbic acid, in either 2% or 0.1% (w/v) glucose to simulate high or reduced-sugar foods, did not reveal reproducible synergies in either of the spoilage yeast species Saccharomyces cerevisiae or Zygosaccharomyces bailii. Potentially promising screen candidates (e.g. lactone parthenolide, ethyl maltol) or a small additional panel of rationally-selected compounds (e.g. benzoic acid) all gave Fractional Inhibitory Concentration Indices (FICI) ≥ 0.5 in combinations with sorbic acid, corroborating absence of synergy in either glucose condition (although FICI values did differ between the glucose conditions). Synergies were not achieved either in a tripartite combination with screen candidates or in a soft-drink formulation as matrix. In previous work with other stressors synergy 'hits' have been comparatively frequent, suggesting that sorbic acid could be unusually resistant to forming synergies with other potential inhibitors and this may relate to the weak acid's known multifactorial inhibitory-actions on cells. The study highlights a challenge in developing appropriate natural product or other chemical combinations applicable to food and beverage preservation.

Evaluation of Sodium Benzoate and Potassium Sorbate Preservative Concentrations in Different Sauce Samples in Urmia, Iran.

Sodium benzoate and potassium sorbate are relatively common preservatives used in a wide range of foods including flavoring products like sauces. The potential health risks arising from these preservatives along with the high-consumption rate of these flavoring products worldwide highlight the importance of the quality and safety assurance of these products. So, this study aimed to evaluate the concentrations of these two common preservatives (i.e., sodium benzoate and potassium sorbate) in different sauce samples, including mayonnaise, salad dressings, Caesar sauce, Italian dressing, Ranch dressing, French dressing, using high-performance liquid chromatography (HPLC) and to compare them with the acceptable level of Codex standard. For this purpose, 49 samples, including three to five samples of each type of different brands of sauce samples, were randomly collected from supermarkets in Urmia, Iran. Based on the results, the mean concentrations ± standard deviation of sodium benzoate and potassium sorbate in the collected samples were found to be 249.9 ± 157 and 158.0 ± 131 ppm, respectively, which were lower than the general standard of the Codex Alimentarius and the European legislation. Due to the importance of hazardous side effects of these preservatives for consumers, regular and accurate evaluation of these preservatives in sauces as highly consumed food products is still recommended for consumer safety.

Simultaneous determination of benzoic acid and sorbic acid in non-alcoholic beverages by a validated HS-GC-MS method with reduced waste.

A headspace gas chromatography-mass spectrometry (HS-GC-MS) method is presented for the simultaneous determination of benzoic acid (BA) and sorbic acid (SoA) in different types of non-alcoholic beverages. Sensitive and reliable results were achieved together with minimising consumption of reagents and samples. Salicylic acid (SalA) was used as internal standard (IS). It was necessary to derivatise BA, SoA and SalA to their methyl esters for HS-GC-MS measurement and extensive optimisation studies for in-vial derivatisation were carried out on the temperature, incubation time, injection time of the loopless HS, as well as on the concentration of sulphuric acid used as a catalyst. Validation studies carried out under optimum conditions after mixing 50 µL of sample and IS solutions with 200 µL of 4.5 M sulphuric acid in 22 mL HS vials revealed that the developed method was both very precise (relative standard deviation < 5%) and accurate (average recovery%: 101.0% for BA and 100.4% for SoA). The validated method was applied to a wide range of beverage types and the results compared with the relevant regulation and product label declarations.