Acetaldehyde metabolism by wine lactic acid bacteria and its oenological implications : a thesis presented in fulfillment of the requirements for the degree of Master of Science in Microbiology at Massey University

Acetaldehyde is one of the most important sensory carbonyl compounds formed during vinification. Excess acetaldehyde can adversely affect the flavour of wine and acetaldehyde plays a role in the colour development of red wines. Excess acetaldehyde is usually masked by the addition of sulphur dioxide (SO2) to the wine (SO2 is also used as an antimicrobial and antioxidant agent in wine and acetaldehyde bound SO2 is less effective in these roles). To date there has been no definitive study of the impact of wine LAB on free and bound acetaldehyde. Therefore, this study investigated the metabolism of free and bound acetaldehyde and its oenological implications. A survey of 11 commercial malolactic starter cultures (mostly Oenococcus oeni strains) showed that 9 out of 11 were able to metabolise acetaldehyde (in a resting state) with the corresponding formation of ethanol and acetic acid as products. SO2 bound acetaldehyde was also metabolised by the two strains tested (Lactobacillus buchneri CUC-3 and Oenococcus oeni MCW). This is the first evidence that LAB can indeed catabolise SO2 bound acetaldehyde, therefore releasing free SO2. During growth Oenococcus oeni EQ54 and Oenococcus oeni VFO were able to metabolise free acetaldehyde in wine at pH 3.3 and pH 3.6. In wine containing SO2 bound acetaldehyde, Oenococcus oeni EQ54 and Oenococcus oeni VFO were able to metabolise SO2 bound acetaldehyde at pH 3.6 after a period of sluggish growth. At pH 3.3 there was no metabolism of SO2 bound acetaldehyde by Oenococcus oeni EQ54 and Oenococcus oeni VFO during the incubation period. Results from growth experiments showed that in broth there was inhibition of growth at 300 mg/L concentration of acetaldehyde for all strains. In wine, no significant inhibition or stimulation of the cultures examined was found at any acetaldehyde concentrations up to 300 mg/L. In a simultaneous resting cell incubation of Saccharomyces bayanus Première Cuvée and Oenococcus oeni Lol11, acetaldehyde produced by the yeast was metabolised by the wine LAB. The metabolism of acetaldehyde by wine LAB is expected to influence wine flavour as small amounts of ethanol and acetic acid are produced and acetaldehyde is removed. This removal of acetaldehyde by wine LAB suggests that less SO2 will need to be added to the wine to mask excess acetaldehyde when malolactic fermentation is performed. Inhibition of wine LAB growth in broth by high levels of acetaldehyde suggests a role for acetaldehyde in stuck or sluggish MLF. Sluggish growth in wine containing SO2 bound acetaldehyde also suggests a possible role of SO2 bound acetaldehyde in stuck and sluggish MLF. This is due to the release of free SO2 through the metabolism of the acetaldehyde moiety of SO2 bound acetaldehyde

Global atmospheric budget of acetaldehyde: 3-D model analysis and constraints from in-situ and satellite observations

We construct a global atmospheric budget for acetaldehyde using a 3-D model of atmospheric chemistry (GEOS-Chem), and use an ensemble of observations to evaluate present understanding of its sources and sinks. Hydrocarbon oxidation provides the largest acetaldehyde source in the model (128 Tg a<sup>−1</sup>, a factor of 4 greater than the previous estimate), with alkanes, alkenes, and ethanol the main precursors. There is also a minor source from isoprene oxidation. We use an updated chemical mechanism for GEOS-Chem, and photochemical acetaldehyde yields are consistent with the Master Chemical Mechanism. We present a new approach to quantifying the acetaldehyde air-sea flux based on the global distribution of light absorption due to colored dissolved organic matter (CDOM) derived from satellite ocean color observations. The resulting net ocean emission is 57 Tg a<sup>−1</sup>, the second largest global source of acetaldehyde. A key uncertainty is the acetaldehyde turnover time in the ocean mixed layer, with quantitative model evaluation over the ocean complicated by known measurement artifacts in clean air. Simulated concentrations in surface air over the ocean generally agree well with aircraft measurements, though the model tends to overestimate the vertical gradient. PAN:NO<sub>x</sub> ratios are well-simulated in the marine boundary layer, providing some support for the modeled ocean source. We introduce the Model of Emissions of Gases and Aerosols from Nature (MEGANv2.1) for acetaldehyde and ethanol and use it to quantify their net flux from living terrestrial plants. Including emissions from decaying plants the total direct acetaldehyde source from the land biosphere is 23 Tg a<sup>−1</sup>. Other terrestrial acetaldehyde sources include biomass burning (3 Tg a<sup>−1</sup>) and anthropogenic emissions (2 Tg a<sup>−1</sup>). Simulated concentrations in the continental boundary layer are generally unbiased and capture the spatial gradients seen in observations over North America, Europe, and tropical South America. However, the model underestimates acetaldehyde levels in urban outflow, suggesting a missing source in polluted air. Ubiquitous high measured concentrations in the free troposphere are not captured by the model, and based on present understanding are not consistent with concurrent measurements of PAN and NO<sub>x</sub>: we find no compelling evidence for a widespread missing acetaldehyde source in the free troposphere. We estimate the current US source of ethanol and acetaldehyde (primary + secondary) at 1.3 Tg a<sup>−1</sup> and 7.8 Tg a<sup>−1</sup>, approximately 60{%} and 480% of the corresponding increases expected for a national transition from gasoline to ethanol fuel

Electron-impact spectroscopy of acetaldehyde

Acetaldehyde has been studied by the technique of low‐energy variable‐angle electron energy‐loss spectroscopy. With this method the low‐lying spin‐forbidden transitions have been located via the behavior of the relative differential cross sections, providing the first identification by this technique of such states in acetaldehyde. High‐lying states were also investigated and some assignments of dipole symmetry‐forbidden/quadrupole symmetry‐allowed excitations were made on the basis of characteristic angular behavior, evident for the asymmetric molecule acetaldehyde just as for the symmetric molecules formaldehyde and acetone. Through a comparison of the acetaldehyde results with those for formaldehyde and acetone the trends in the allowed and forbidden transition energies were examined as a function of methyl substitution and found to be relatively linear

Quantifying Acetaldehyde in Astronomical Ices and Laboratory Analogues: IR Spectra, Intensities, 13C Shifts, and Radiation Chemistry

Acetaldehyde is of interest to astrochemists for its relevance to both interstellar and cometary chemistry, but little infrared (IR) spectral data have been published for the solid phases of this compound. Here we present IR spectra of three forms of solid acetaldehyde, with spectra for one form being published for the first time. Direct measurements of band strengths and absorption coefficients also are reported for the first time for amorphous aldehyde, the form of greatest interest for astrochemical work. An acetaldehyde band strength at 1350 cm1 that has been used as a reference for about 20 yr is seen to be in error by about 80 per cent when compared to the direct measurements presented here. Spectra and peak positions also are presented for H13C(O)13CH3, and then used for the first identification of ketene as a radiation product of solid acetaldehyde

The effect of flooding on the exchange of the volatile C₂-compounds ethanol, acetaldehyde and acetic acid between leaves of Amazonian floodplain tree species and the atmosphere

The effect of root inundation on the leaf emissions of ethanol, acetaldehyde and acetic acid in relation to assimilation and transpiration was investigated with 2–3 years old tree seedlings of four Amazonian floodplain species by applying dynamic cuvette systems under greenhouse conditions. Emissions were monitored over a period of several days of inundation using a combination of Proton Transfer Reaction Mass Spectrometry (PTR-MS) and conventional techniques (HPLC, ion chromatography). Under non-flooded conditions, none of the species exhibited measurable emissions of any of the compounds, but rather low deposition of acetaldehyde and acetic acid was observed instead. Tree species specific variations in deposition velocities were largely due to variations in stomatal conductance. Flooding of the roots resulted in leaf emissions of ethanol and acetaldehyde by all species, while emissions of acetic acid were only observed from the species exhibiting the highest ethanol and acetaldehyde emission rates. All three compounds showed a similar diurnal emission profile, each displaying an emission burst in the morning, followed by a decline in the evening. This concurrent behavior supports the conclusion, that all three compounds emitted by the leaves are derived from ethanol produced in the roots by alcoholic fermentation, transported to the leaves with the transpiration stream and finally partly converted to acetaldehyde and acetic acid by enzymatic processes. Co-emissions and peaking in the early morning suggest that root ethanol, after transportation with the transpiration stream to the leaves and enzymatic oxidation to acetaldehyde and acetate, is the metabolic precursor for all compounds emitted, though we can not totally exclude other production pathways. Emission rates substantially varied among tree species, with maxima differing by up to two orders of magnitude (25–1700 nmol m−2 min−1 for ethanol and 5–500 nmol m−2 min−1 for acetaldehyde). Acetic acid emissions reached 12 nmol m−2 min−1. The observed differences in emission rates between the tree species are discussed with respect to their root adaptive strategies to tolerate long term flooding, providing an indirect line of evidence that the root ethanol production is a major factor determining the foliar emissions. Species which develop morphological root structures allowing for enhanced root aeration produced less ethanol and showed much lower emissions compared to species which lack gas transporting systems, and respond to flooding with substantially enhanced fermentation rates and a non-trivial loss of carbon to the atmosphere. The pronounced differences in the relative emissions of ethanol to acetaldehyde and acetic acid between the tree species indicate that not only the ethanol production in the roots but also the metabolic conversion in the leaf is an important factor determining the release of these compounds to the atmosphere

An experimental study of adsorption interference in binary mixtures flowing through activated carbon

The isothermal transmission through activated carbon adsorber beds at 25 C of acetaldehyde-propane and acetylene-ethane mixtures in a helium carrier gas was measured. The inlet concentration of each component was in the range between 10 ppm and 500 ppm. The constant inlet volumetric flow rate was controlled at 200 cc (STP)/min in the acetaldehyde-propane experiments and at 50 cc (STP)/min in the acetaldehyde-ethane experiments. Comparison of experimental results with the corresponding single-component experiments under similar conditions reveals interference phenomena between the components of the mixtures as evidenced by changes in both the adsorption capacity and the dispersion number. Propane was found to displace acetaldehyde from the adsorbed state. The outlet concentration profiles of propane in the binary mixtures tend to become more diffuse than the corresponding concentration profiles of the one-component experiments. Similar features were observed with mixtures of acetylene and ethane; however, the displacement of acetylene by ethane is less pronounced

Preventing hereditary cancers caused by opportunistic carcinogens

Objectives
Previous studies reported inherited BRCA1/2 deficits appear to cause cancer by impairing normal protective responses to some carcinogens. Opportunistic carcinogens can exploit these deficits by causing chronic inflammation, constant cell death and replacement in a mutagenic environment, DNA crosslinking or double strand breaks. Some of the resulting cancers may be prevented if BRCA1/2 specific carcinogens are identified.
Methods
The literature was systematically searched for carcinogens capable of exploiting deficits in BRCA1/2 pathways. Search criteria were common exposure, available information, required BRCA1/2 pathway repairs, increased risks for any cancer, and effects on stem cells.
Results
Formaldehyde and acetaldehyde are closely related carcinogens and common pollutants that are everywhere. Alcohol metabolism also produces acetaldehyde. High levels of either carcinogen overwhelm normal detoxification systems, cause inflammation, inhibit DNA repair and produce DNA cross links as critical carcinogenic lesions. Searching model system studies revealed both carcinogens activate stem cells, BRCA1/2 pathways and connected BRCA1/2 pathways to myeloid leukemia. For example, the BRCA1-BARD1 complex is required for proper nucleophosmin functions. Nucleophosmin prevents a major subset of acute myeloid leukemia (AML). Next, these concepts were independently tested against risks for myeloid leukemia. Epidemiologic results showed that BRCA2 gene defects inherited on both chromosomes increased risks so dramatically that AML occurs in most children. Using data from 14 studies, known/potential heterozygous BRCA1/BRCA2 mutations increased risks for myeloid leukemias by at least 3 fold in 7 studies and by at least 50% in 12.
Acetaldehyde occurs in breast milk. In model studies, excessive acetaldehyde/alcohol exposure affects estrogen metabolism and stimulates alternate alcohol detoxification pathways.
These pathways can cause DNA cross linking by releasing oxygen species and activating procarcinogens. Acetaldehyde in rats’ drinking water increased incidence of leukemias, lymphomas,pancreatic cancers and fibroadenomas. Human epidemiologic studies showed increased premenopausal breast cancer risks associated with persistent/high acetaldehyde levels related to alcohol metabolism genotype.
Conclusions
Although it is difficult to prove direct causation, BRCA1/2 mutation carriers may reduce cancer risks by avoiding excessive formaldehyde and acetaldehyd

Reverse chemical ecology approach for the identification of an oviposition attractant for Culex quinquefasciatus.

Pheromones and other semiochemicals play a crucial role in today's integrated pest and vector management strategies. These semiochemicals are typically discovered by bioassay-guided approaches. Here, we applied a reverse chemical ecology approach; that is, we used olfactory proteins to lead us to putative semiochemicals. Specifically, we used 7 of the top 10 odorant receptors (ORs) most expressed in the antennae of the southern house mosquito, Culex quinquefasciatus, and which are yet to be deorphanized. We expressed these receptors in the Xenopus oocyte recording system and challenged them with a panel of 230 odorants, including physiologically and behaviorally active compounds. Six of the ORs were silent either because they are not functional or a key odorant was missing. CquiOR36, which showed the highest transcript levels of all OR genes in female antennae, was also silent to all odorants in the tested panel, but yielded robust responses when it was accidentally challenged with an old sample of nonanal in ethanol. After confirming that fresh samples were inactive and through a careful investigation of all possible "contaminants" in the old nonanal samples, we identified the active ligand as acetaldehyde. That acetaldehyde is activating CquiOR36 was further confirmed by electroantennogram recordings from antennae of fruit flies engineered to carry CquiOR36. Antennae of female mosquitoes also responded to acetaldehyde. Cage oviposition and dual-choice assays demonstrated that acetaldehyde is an oviposition attractant in a wide range of concentrations and thus of potential practical applications

Evaluation of the acetaldehyde production and degradation potential of 26 enological Saccharomyces and non- Saccharomyces yeast strains in a resting cell model system

Acetaldehyde is relevant for wine aroma, wine color, and microbiological stability. Yeast are known to play a crucial role in production and utilization of acetaldehyde during fermentations but comparative quantitative data are scarce. This research evaluated the acetaldehyde metabolism of 26 yeast strains, including commercial Saccharomyces and non-Saccharomyces, in a reproducible resting cell model system. Acetaldehyde kinetics and peak values were highly genus, species, and strain dependent. Peak acetaldehyde values varied from 2.2 to 189.4mgl−1 and correlated well (r 2=0.92) with the acetaldehyde production yield coefficients that ranged from 0.4 to 42mg acetaldehyde per g of glucose in absence of SO2. S. pombe showed the highest acetaldehyde production yield coefficients and peak values. All other non-Saccharomyces species produced significantly less acetaldehyde than the S. cerevisiae strains and were less affected by SO2 additions. All yeast strains could degrade acetaldehyde as sole substrate, but the acetaldehyde degradation rates did not correlate with acetaldehyde peak values or acetaldehyde production yield coefficients in incubations with glucose as sole substrat

Bacteria in the airways of patients with cystic fibrosis are genetically capable of producing VOCs in breath.

Breath contains hundreds of volatile organic compounds (VOCs), the composition of which is altered in a wide variety of diseases. Bacteria are implicated in the formation of VOCs, but the biochemical mechanisms that lead to the formation of breath VOCs remain largely hypothetical. We hypothesized that bacterial DNA fragments in sputum of CF patients could be sequenced to identify whether the bacteria present were capable of producing VOCs found in the breath of these patients. Breath from seven patients with cystic fibrosis was sampled and analyzed by gas-chromatography and mass-spectrometry. Sputum samples were also collected and microbial DNA was isolated. Metagenomic sequencing was performed and the DNA fragments were compared to a reference database with genes that are linked to the metabolism of acetaldehyde, ethanol and methanol in the KEGG database. Bacteria in the genera Escherichia, Lactococcus, Pseudomonas, Rothia and Streptococcus were found to have the genetic potential to produce acetaldehyde and ethanol. Only DNA sequences from Lactococcus were implicated in the formation of acetaldehyde from acetate through aldehyde dehydrogenase family 9 member A1 (K00149). Escherichia was found to be genetically capable of producing ethanol in all patients, whilst there was considerable heterogeneity between patients for the other genera. The ethanol concentration in breath positively correlated with the amount of Escherichia found in sputum (Spearman rho  =  0.85,  P  =  0.015). Rothia showed the most versatile genetic potential for producing methanol. To conclude, bacterial DNA fragments in sputum of CF patients can be linked to enzymes implicated in the production of ethanol, acetaldehyde and methanol, which are VOCs that are predictive of respiratory tract colonization and/or infection. This supports that the lung microbiome can produce VOCs directly