A new HS-SPME-GC-MS analytical method to identify and quantify compounds responsible for changes in the volatile profile in five types of meat products during aerobic storage at 4 °C.

Nowadays, it is important to monitor the freshness of meat during storage to protect consumers' health. Volatile organic compounds (VOCs) are responsible for odour and taste of food, and they give an indication about meat quality and freshness. This study had the aim to seek and select potential new markers of meat spoilage through a semi-quantitative analysis in five types of meat (beef, raw and baked ham, pork sausage and chicken) and then to develop a new quantitative analytical method to detect and quantify potential markers on five types of meat simultaneously. Firstly, a new headspace-solid phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) method was developed to evaluate the volatile profile of five types of meat, preserved at 4 °C for 5 days. Among the 40 compounds identified, 15 were chosen and selected as potential shelf-life markers on the basis of their presence in most of meat samples or/and for their constant increasing/decreasing trend within the sample. Afterwards, a quantitative HS-SPME-GC-MS analytical method was developed to confirm which VOCs can be considered markers of shelf-life for these meat products, stored at 4 °C for 12 days. Some of the compounds analyzed attracted attention as they can be considered markers of shelf-life for at least 4 types of meat: 1-butanol, 3-methylbutanol, 1-hexanol, 2-nonanone, nonanal, 1-octen-3-ol and linalool. In conclusion, in this study a new quantitative HS-SPME-GC-MS analytical method to quantity 15 VOCs in five types of meat was developed and it was demonstrated that some of the compounds quantified can be considered markers of shelf-life for some of the meat products analyzed.

Decoding the chemical language of Suillus fungi: genome mining and untargeted metabolomics uncover terpene chemical diversity.

Ectomycorrhizal fungi establish mutually beneficial relationships with trees, trading nutrients for carbon. Suillus are ectomycorrhizal fungi that are critical to the health of boreal and temperate forest ecosystems. Comparative genomics has identified a high number of non-ribosomal peptide synthetase and terpene biosynthetic gene clusters (BGC) potentially involved in fungal competition and communication. However, the functionality of these BGCs is not known. This study employed co-culture techniques to activate BGC expression and then used metabolomics to investigate the diversity of metabolic products produced by three Suillus species (Suillus hirtellus EM16, Suillus decipiens EM49, and Suillus cothurnatus VC1858), core members of the pine microbiome. After 28 days of growth on solid media, liquid chromatography-tandem mass spectrometry identified a diverse range of extracellular metabolites (exometabolites) along the interaction zone between Suillus co-cultures. Prenol lipids were among the most abundant chemical classes. Out of the 62 unique terpene BGCs predicted by genome mining, 41 putative prenol lipids (includes 37 putative terpenes) were identified across the three Suillus species using metabolomics. Notably, some terpenes were significantly more abundant in co-culture conditions. For example, we identified a metabolite matching to isomers isopimaric acid, sandaracopimaric acid, and abietic acid, which can be found in pine resin and play important roles in host defense mechanisms and Suillus spore germination. This research highlights the importance of combining genomics and metabolomics to advance our understanding of the chemical diversity underpinning fungal signaling and communication.IMPORTANCEUsing a combination of genomics and metabolomics, this study's findings offer new insights into the chemical diversity of Suillus fungi, which serve a critical role in forest ecosystems.

Metabolomic insights into the browning inhibition of fresh-cut apple by hydrogen sulfide.

Hydrogen sulfide (H2S) is known to effectively inhibit the browning of fresh-cut apples, but the mechanism at a metabolic level remains unclear. Herein, non-targeted metabolomics was used to analyze metabolic changes in surface and internal tissues of fresh-cut apple after H2S treatment. The results showed that prenol lipids were the most up-accumulated differential metabolites in both surface and inner tissue of fresh-cut apple during browning process, which significantly down-accumulated by H2S treatment. H2S treatment reduced the consumption of amino acid in surface tissue. Regarding inner tissue, H2S activated defense response through accumulation of lysophospholipid signaling and induced the biosynthesis of phenolic compounds. We therefore propose that H2S inhibited the surface browning of fresh-cut apple by reducing the accumulation of prenol lipids, directly delaying amino acid consumption in surface tissue and indirectly regulating defense response in inner tissue, which provides fundamental insights into browning inhibition mechanisms by H2S.

Excess Enthalpies Analysis of Biofuel Components: Sunflower Oil-Alcohols Systems.

This study addresses the pressing issues of energy production and consumption, in line with global sustainable development goals. Focusing on the potential of alcohols as "green" alternatives to traditional fossil fuels, especially in biofuel applications, we investigate the thermochemical properties of three alcohols (n-propanol, n-butanol, n-pentanol) blended with sunflower oil. The calorimetric analysis allows for the experimental determination of excess enthalpies in pseudo-binary mixtures at 303.15 K, revealing similarities in the trends of the curves (dependence on concentrations) but with different values for the excess enthalpies for each mixture. Despite the structural differences of the alcohols studied, the molar excess enthalpy values exhibit uniformity, suggesting consistent mixing behavior. The peak values of excess enthalpies for systems with sunflower oil and n-propanol, n-butanol and n-pentanol are, respectively, 3255.2 J/mole, 3297.4 J/mole and 3150.1 J/mole. Both the NRTL and Redlich-Kister equations show satisfactory agreement with the obtained values.

Volatilomic signatures of different strains of Helicobacter pylori.

BACKGROUND: Helicobacter pylori (H. pylori) infection is the most extensively studied risk factor for gastric cancer. As with any bacteria, H. pylori will release distinctive odors that result from an emission of volatile metabolic byproducts in unique combinations and proportions. Effectively capturing and identifying these volatiles can pave the way for the development of innovative and non-invasive diagnostic methods for determining infection. Here we characterize the H. pylori volatilomic signature, pinpoint potential biomarkers of its presence, and evaluate the variability of volatilomic signatures between different H. pylori isolates. MATERIALS AND METHODS: Using needle trap extraction, volatiles in the headspace above H. pylori cultures were collected and, following thermal desorption at 290°C in a splitless mode, were analyzed using gas chromatography-mass spectrometry. The resulting volatilomic signatures of H. pylori cultures were compared to those obtained from an analysis of the volatiles in the headspace above the cultivating medium only. RESULTS: Amongst the volatiles detected, 21 showed consistent differences between the bacteria cultures and the cultivation medium, with 11 compounds being elevated and 10 showing decreased levels in the culture's headspace. The 11 elevated volatiles are four ketones (2-pentanone, 5-methyl-3-heptanone, 2-heptanone, and 2-nonanone), three alcohols (2-methyl-1-propanol, 3-methyl-1-butanol, and 1 butanol), one aromatic (styrene), one aldehyde (2-ethyl-hexanal), one hydrocarbon (n-octane), and one sulfur compound (dimethyl disulfide). The 10 volatiles with lower levels in the headspace of the cultures are four aldehydes (2-methylpropanal, benzaldehyde, 3-methylbutanal, and butanal), two heterocyclic compounds (2-ethylfuran and 2-pentylfuran), one ketone (2-butanone), one aromatic (benzene), one alcohol (2-butanol) and bromodichloromethane. Of the volatile species showing increased levels, the highest emissions are found to be for 3-methyl-1-butanol, 1-butanol and dimethyl disulfide. Qualitative variations in their emissions from the different isolates was observed. CONCLUSIONS: The volatiles emitted by H. pylori provide a characteristic volatilome signature that has the potential of being developed as a tool for monitoring infections caused by this pathogen. Furthermore, using the volatilome signature, we are able to differentiate different isolates of H. pylori. However, the volatiles also represent potential confounders for the recognition of gastric cancer volatile markers.

A Ribulose-5-phosphate Shunt from the Calvin-Benson Cycle to Methylerythritol Phosphate Pathway for Enhancing Photosynthetic Terpenoid Production.

Cyanobacteria are attractive hosts for photosynthetic terpenoid production, using CO2 as the sole carbon source. Although the methylerythritol phosphate (MEP) pathway is superior to the mevalonate pathway for cyanobacterial terpenoid synthesis, the first reaction of the MEP pathway, which is catalyzed by 1-deoxy-d-xylulose-5-phosphate (DXP) synthase, involves complex regulation and carbon loss. Here, we constructed a direct route linking ribulose-5-phosphate (Ru5P) in the Calvin-Benson (CB) cycle with DXP in the MEP pathway in a cyanobacterium to increase the terpenoid yield from CO2 and bypass the DXS-targeted regulations. By employing the adaptive laboratory evolution, we identified new RibB variants including RibB 90-92del with a high activity of synthesizing DXP from Ru5P. These RibB variants were introduced into Synechococcus elongatus, resulting in the significantly increased photosynthetic production of isopentenol. The 13C tracer experiments demonstrated a direct carbon flow from Ru5P in the CB cycle to the MEP pathway; thus, this direct route was denoted as the Ru5P shunt. The strain harboring the Ru5P shunt produced 105.2 mg L-1 of isopentenol with an average rate of 17.5 mg L-1 d-1 under continuous light conditions, which is higher than those ever reported for five-carbon alcohol production by photoautotrophic microorganisms. Utilization of the Ru5P shunt in cyanobacterial cells also improved the pinene production, which demonstrates that this shunt can be used to enhance the photosynthetic production of diverse terpenoids.

Comparative study of two indoor microbial volatile pollutants, 2-Methyl-1-butanol and 3-Methyl-1-butanol, on growth and antioxidant system of rice (Oryza sativa) seedlings.

2-Methyl-1-butanol (2MB) and 3-Methyl-1-butanol (3MB) are microbial volatile organic compounds (VOCs) and found in indoor air. Here, we applied rice as a bioindicator to investigate the effects of these indoor microbial volatile pollutants. A remarkable decrease in germination percentage, shoot and root elongation, as well as lateral root numbers were observed in 3MB. Furthermore, ROS production increased by 2MB and 3MB, suggesting that pentanol isomers could induce cytotoxicity in rice seedlings. The enhancement of peroxidase (POD) and catalase (CAT) activity provided evidence that pentanol isomers activated the enzymatic antioxidant scavenging systems, with a more significant effect observed in 3MB. Furthermore, 3MB induced higher activity levels of glutathione (GSH), oxidized glutathione (GSSG), and the GSH/GSSG ratio in rice compared to the levels induced by 2MB. Additionally, qRT-PCR analysis showed more up-regulation in the expression of glutaredoxins (GRXs), peroxiredoxins (PRXs), thioredoxins (TRXs), and glutathione S-transferases (GSTUs) genes in 3MB. Taking the impacts of pentanol isomers together, the present study suggests that 3MB exhibits more cytotoxic than 2MB, as such has critical effects on germination and the early seedling stage of rice. Our results provide molecular insights into how isomeric indoor microbial volatile pollutants affect plant growth through airborne signals.

Volatile Organic Compounds (VOCs) Produced by Levilactobacillus brevis WLP672 Fermentation in Defined Media Supplemented with Different Amino Acids.

Fermentation by lactic acid bacteria (LAB) is a promising approach to meet the increasing demand for meat or dairy plant-based analogues with realistic flavours. However, a detailed understanding of the impact of the substrate, fermentation conditions, and bacterial strains on the volatile organic compounds (VOCs) produced during fermentation is lacking. As a first step, the current study used a defined medium (DM) supplemented with the amino acids L-leucine (Leu), L-isoleucine (Ile), L-phenylalanine (Phe), L-threonine (Thr), L-methionine (Met), or L-glutamic acid (Glu) separately or combined to determine their impact on the VOCs produced by Levilactobacillus brevis WLP672 (LB672). VOCs were measured using headspace solid-phase microextraction (HS-SPME) gas chromatography-mass spectrometry (GC-MS). VOCs associated with the specific amino acids added included: benzaldehyde, phenylethyl alcohol, and benzyl alcohol with added Phe; methanethiol, methional, and dimethyl disulphide with added Met; 3-methyl butanol with added Leu; and 2-methyl butanol with added Ile. This research demonstrated that fermentation by LB672 of a DM supplemented with different amino acids separately or combined resulted in the formation of a range of dairy- and meat-related VOCs and provides information on how plant-based fermentations could be manipulated to generate desirable flavours.

Novel approach for corn straw biorefineries: Production of xylooligosaccharides, lignin and ethanol by nicotinic acid hydrolysis and pentanol pretreatment.

The productive separation and conversion of corn straw offers significant prospects for the economic viability of biorefineries centered on straw resources. In this work, a graded utilization method was proposed to produce xylo-oligosaccharides (XOS), ethanol and lignin from corn straw by nicotinic acid (NA) hydrolysis and water/pentanol pretreatment. A XOS yield of 52.6 % was achieved under optimized conditions of 100 mM NA, 170 °C and 30 min. The solid residue was directly treated with water/pentanol, achieving a lignin removal rate of 79.7 %, and the total XOS yield was improved to 62.6 %. The lignin recovered from pentanol had a high purity of 97.6 %, with high phenolic OH content. Simultaneous saccharification and fermentation of final residue resulted in an ethanol yield of 92.0 %, which yielded 55.3 g/L ethanol. Thus, NA hydrolysis and water/pentanol pretreatment provided an efficient, environmentally friendly approach to fractionate corn straw for the co-production of XOS, ethanol, and lignin.

Signaling pathways involved in microbial indoor air pollutant 3-methyl-1-butanol in the induction of stomatal closure in Arabidopsis.

Indoor air pollution is a global problem and one of the main stress factors that has negative effects on plant and human health. 3-methyl-1-butanol (3MB), an indoor air pollutant, is a microbial volatile organic compound (mVOC) commonly found in damp indoor dwellings. In this study, we reported that 1 mg/L of 3MB can elicit a significant reduction in the stomatal aperture ratio in Arabidopsis and tobacco. Our results also showed that 3MB enhances the reactive oxygen species (ROS) production in guard cells of wild-type Arabidopsis after 24 h exposure. Further investigation of 24 h 3MB fumigation of rbohD, the1-1, mkk1, mkk3, and nced3 mutants revealed that ROS production, cell wall integrity, MAPK kinases cascade, and phytohormone abscisic acid are all involved in the process of 3MB-induced stomatal. Our findings proposed a mechanism by which 3MB regulates stomatal closure in Arabidopsis. Understanding the mechanisms by which microbial indoor air pollutant induces stomatal closure is critical for modulating the intake of harmful gases from indoor environments into leaves. Investigations into how stomata respond to the indoor mVOC 3MB will shed light on the plant's "self-defense" system responding to indoor air pollution.

Volatiles from nutritional fungal symbiont influence the attraction of Anisandrus maiche (Coleoptera: Curculionidae) to ethanol-baited traps.

Anisandrus maiche Stark (Coleoptera: Curculionidae: Scolytinae) is a non-native ambrosia beetle from central Asia that has been spreading throughout the eastern United States since 2005. Preferred hosts of A. maiche are not well characterized within its currently invaded range, but it is established in managed and natural forests throughout Indiana. Current monitoring and detection efforts for this beetle rely on ethanol-baited traps, but fungal volatiles may alter the attraction of A. maiche to ethanol. In this study, we conducted trapping experiments in Indiana to determine the extent to which a suite of common fungal alcohols influences the response of A. maiche to ethanol-baited traps. We then evaluated isoamyl and isobutyl alcohol as potential attractants for A. maiche and their ability to enhance attraction to ethanol. Lastly, we used SPME-GC-MS to identify volatiles from Ambrosiella cleistominuta (Mayers & Harr.), the fungal symbiont of A. maiche, grown for 7 and 14 days on malt extract agar. Benzyl alcohol, isobutyl alcohol, hexanol, methyl phenylacetate, phenethyl alcohol, and piperitone reduced the attraction of A. maiche to ethanol-baited traps in the field. Moreover, adding methyl benzoate and isoamyl alcohol individually to ethanol-baited traps did not further increase A. maiche capture. When paired with ethanol, isoamyl alcohol repelled beetles in the early flight period but did not significantly increase trap capture during the fall flight. These results represent a first step in understanding the role of fungal volatiles in the colonization behavior of A. maiche and may ultimately inform management strategies for this species.

Biphasic pretreatment excels over conventional sulfuric acid in pinewood biorefinery: An environmental analysis.

This study assesses the environmental impact of pine chip-based biorefinery processes, focusing on bioethanol, xylonic acid, and lignin production. A cradle-to-gate Life Cycle Assessment (LCA) is employed, comparing a novel biphasic pretreatment method (p-toluenesulfonic acid (TsOH)/pentanol, Sc-1) with conventional sulfuric acid pretreatment (H2SO4, Sc-2). The analysis spans biomass handling, pretreatment, enzymatic hydrolysis, yeast fermentation, and distillation. Sc-1 yielded an environmental impact of 1.45E+01 kPt, predominantly affecting human health (96.55%), followed by ecosystems (3.07%) and resources (0.38%). Bioethanol, xylonic acid, and lignin contributed 32.61%, 29.28%, and 38.11% to the total environmental burdens, respectively. Sc-2 resulted in an environmental burden of 1.64E+01 kPt, with a primary impact on human health (96.56%) and smaller roles for ecosystems (3.07%) and resources (0.38%). Bioethanol, xylonic acid, and lignin contributed differently at 22.59%, 12.5%, and 64.91%, respectively. Electricity generation was predominant in both scenarios, accounting for 99.05% of the environmental impact, primarily driven by its extensive usage in biomass handling and pretreatment processes. Sc-1 demonstrated a 13.05% lower environmental impact than Sc-2 due to decreased electricity consumption and increased bioethanol and xylonic acid outputs. This study highlights the pivotal role of pretreatment methods in wood-based biorefineries and underscores the urgency of sustainable alternatives like TsOH/pentanol. Additionally, adopting greener electricity generation, advanced technologies, and process optimization are crucial for reducing the environmental footprint of waste-based biorefineries while preserving valuable bioproduct production.

Beyond the MEP Pathway: A novel kinase required for prenol utilization by malaria parasites.

A proposed treatment for malaria is a combination of fosmidomycin and clindamycin. Both compounds inhibit the methylerythritol 4-phosphate (MEP) pathway, the parasitic source of farnesyl and geranylgeranyl pyrophosphate (FPP and GGPP, respectively). Both FPP and GGPP are crucial for the biosynthesis of several essential metabolites such as ubiquinone and dolichol, as well as for protein prenylation. Dietary prenols, such as farnesol (FOH) and geranylgeraniol (GGOH), can rescue parasites from MEP inhibitors, suggesting the existence of a missing pathway for prenol salvage via phosphorylation. In this study, we identified a gene in the genome of P. falciparum, encoding a transmembrane prenol kinase (PolK) involved in the salvage of FOH and GGOH. The enzyme was expressed in Saccharomyces cerevisiae, and its FOH/GGOH kinase activities were experimentally validated. Furthermore, conditional knockout parasites (Δ-PolK) were created to investigate the biological importance of the FOH/GGOH salvage pathway. Δ-PolK parasites were viable but displayed increased susceptibility to fosmidomycin. Their sensitivity to MEP inhibitors could not be rescued by adding prenols. Additionally, Δ-PolK parasites lost their capability to utilize prenols for protein prenylation. Experiments using culture medium supplemented with whole/delipidated human plasma in transgenic parasites revealed that human plasma has components that can diminish the effectiveness of fosmidomycin. Mass spectrometry tests indicated that both bovine supplements used in culture and human plasma contain GGOH. These findings suggest that the FOH/GGOH salvage pathway might offer an alternate source of isoprenoids for malaria parasites when de novo biosynthesis is inhibited. This study also identifies a novel kind of enzyme related to isoprenoid metabolism.

Chemical basis of microbiome preference in the nematode C. elegans.

Animals are exposed to many microbes in their environment, some of which have been shown to colonize various tissues including the intestine. The composition of the intestinal microbiota affects many aspects of the host's physiology and health. Despite this, very little is known about whether host behavior contributes to the colonization. We approach this question in the nematode C. elegans, which feeds on bacteria and also harbors an intestinal microbiome. We examined the behavior of C. elegans towards CeMbio, a simplified microbiome consisting of twelve strains that represent the bacteria found in the animal's natural environment. We observed that C. elegans raised on E. coli shows a strong preference for three members of CeMbio (Lelliottia amnigena JUb66, Enterobacter hormaechei CEent1, and Pantoea nemavictus BIGb0393) compared to E. coli. Previously, these three bacterial strains have been shown to support faster C. elegans development time than E. coli OP50 and are low colonizers compared to eight other members of CeMbio. We then used gas chromatography coupled to mass spectrometry to identify that these three bacteria release isoamyl alcohol, a previously described C. elegans chemoattractant. We suggest that C. elegans seeks bacteria that release isoamyl alcohol and support faster growth.

THI3 contributes to isoamyl alcohol biosynthesis through thiamine diphosphate homeostasis.

Isoamyl alcohol is a precursor of isoamyl acetate, an aromatic compound that imparts the ginjo aroma to sake. The isoamyl alcohol biosynthesis pathway in yeasts involves the genes PDC1, PDC5, PDC6, ARO10, and THI3 encoding enzymes that decarboxylate α-ketoisocaproic acid to isovaleraldehyde. Among these genes, THI3 is the main gene involved in isoamyl alcohol biosynthesis. Decreased production of isoamyl alcohol has been reported in yeast strains with disrupted THI3 (Δthi3). However, it has also been reported that high THI3 expression did not enhance decarboxylase activity. Therefore, the involvement of THI3 in isoamyl alcohol biosynthesis remains unclear. In this study, we investigated the role of THI3 in isoamyl alcohol biosynthesis. While reproducing previous reports of reduced isoamyl alcohol production by the Δthi3 strain, we observed that the decrease in isoamyl alcohol production occurred only at low yeast nitrogen base concentrations in the medium. Upon investigating individual yeast nitrogen base components, we found that the isoamyl alcohol production by the Δthi3 strain reduced when thiamine concentrations in the medium were low. Under low-thiamine conditions, both thiamine and thiamine diphosphate (TPP) levels decreased in Δthi3 cells. We also found that the decarboxylase activity of cell-free extracts of the Δthi3 strain cultured in a low-thiamine medium was lower than that of the wild-type strain, but was restored to the level of the wild-type strain when TPP was added. These results indicate that the loss of THI3 lowers the supply of TPP, a cofactor for decarboxylases, resulting in decreased isoamyl alcohol production.

Amyl Acetate and Isoamyl Acetate.

The Expert Panel for Cosmetic Ingredient Safety reviewed newly available studies since their original assessment in 1988, along with updated information regarding product types and concentrations of use, and confirmed that Amyl Acetate and Isoamyl Acetate are safe as cosmetic ingredients in the practices of use and concentration as described in this report.

Rapid construction of Escherichia coli chassis with genome multi-position integration of isopentenol utilization pathway for efficient and stable terpenoid accumulation.

The isopentenol utilization pathway (IUP) is potential in terpenoids synthesis. This study aimed to construct IUP-employed Escherichia coli chassis for stably synthesizing terpenoids. As to effectiveness, promotor engineering strategy was employed to regulate IUP expression level, while ribosome-binding site (RBS) library of the key enzyme was constructed for screening the optimal RBS, followed by optimization of concentration of inducer and substrates, the titer of reporting production, lycopene, from 0.087 to 8.67 mg OD600 -1 . As about stability, the IUP expression cassette was integrated into the genome through transposition tool based on CRISPR-associated transposases. Results showed that the strain with 13 copies produced 1.78-fold lycopene titer that of the controlled strain with IUP-harbored plasmid, and it exhibited stable expression after ten successions while the plasmid loss was observed in the controlled strain in the 3rd succession. This strategy provides valuable information for rapid construction of highly effective and stable chassis employing IUP for terpenoids production.