Free Radicals in Little Cigar Mainstream Smoke and the Potential Influence of Flavoring Chemicals on Free Radical Production.

Implementation of the Tobacco Control Act in 2009 banned characterizing flavors in cigarettes (except menthol and tobacco), but substitution has occurred by the continued availability of alternative flavored products (i.e., flavored little cigars). Little is known about how flavorants in noncigarette tobacco products impact human health. Thus, we investigated the impact of flavorants on free radical production in the mainstream smoke of little cigars. Gas- and particulate-phase free radical yields in mainstream smoke generated from 12 commercial little cigar brands and research little cigars and cigarettes were measured via electron paramagnetic resonance spectroscopy using the International Organization of Standardization (ISO) smoking protocol. Flavorants were extracted from unsmoked little cigars and analyzed by gas chromatography-mass spectroscopy. Gas- and particulate-phase radical yields from little cigars ranged from 13.5 to 97.6 and 0.453-1.175 nmol/unit, respectively. Comparatively, research cigarettes yielded an average of 4.9 nmol gas-phase radicals/unit and 0.292 nmol particulate-phase radicals/unit. From the products, 66 flavorants were identified, with each brand containing 4-24 individual flavorants. The free radical content was strongly correlated with the number of flavorants present in each cigar (r = 0.74, p = 0.01), indicating that highly flavored little cigars may produce higher levels of toxic free radicals. The presence of the flavorant ethyl methylphenylglycidate (strawberry) was associated with >2-fold higher levels of GP radicals (p = 0.001). Our results show that free radical delivery from little cigars is greater than that from research cigarettes and provide empirical evidence for the harmfulness of flavored tobacco products. Additionally, it demonstrates that flavorants present in combustible tobacco products can influence the levels of free radicals produced. Therefore, future tobacco product standards should consider little cigars.

Unburned Tobacco Cigarette Smoke Alters Rat Ultrastructural Lung Airways and DNA.

INTRODUCTION: Recently, the Food and Drug Administration authorized the marketing of IQOS Tobacco Heating System as a Modified Risk Tobacco Product based on an electronic heat-not-burn technology that purports to reduce the risk. METHODS: Sprague-Dawley rats were exposed in a whole-body mode to IQOS aerosol for 4 weeks. We performed the chemical characterization of IQOS mainstream and we studied the ultrastructural changes in trachea and lung parenchyma of rats exposed to IQOS stick mainstream and tissue pro-inflammatory markers. We investigated the reactive oxygen species amount along with the markers of tissue and DNA oxidative damage. Moreover, we tested the putative genotoxicity of IQOS mainstream through Ames and alkaline Comet mutagenicity assays. RESULTS: Here, we identified irritating and carcinogenic compounds including aldehydes and polycyclic aromatic hydrocarbons in the IQOS mainstream as sign of incomplete combustion and degradation of tobacco, that lead to severe remodelling of smaller and largest rat airways. We demonstrated that IQOS mainstream induces lung enzymes that activate carcinogens, increases tissue reactive radical concentration; promotes oxidative DNA breaks and gene level DNA damage; and stimulates mitogen activated protein kinase pathway which is involved in the conventional tobacco smoke-induced cancer progression. CONCLUSIONS: Collectively, our findings reveal that IQOS causes grave lung damage and promotes factors that increase cancer risk. IMPLICATIONS: IQOS has been proposed as a safer alternative to conventional cigarettes, due to depressed concentration of various harmful constituents typical of traditional tobacco smoke. However, its lower health risks to consumers have yet to be determined. Our findings confirm that IQOS mainstream contains pyrolysis and thermogenic degradation by-products, the same harmful constituents of traditional cigarette smoke, and, for the first time, we show that it causes grave lung damage and promotes factors that increase cancer risk in the animal model.

Gliadin Sequestration as a Novel Therapy for Celiac Disease: A Prospective Application for Polyphenols.

Celiac disease is an autoimmune disorder characterized by a heightened immune response to gluten proteins in the diet, leading to gastrointestinal symptoms and mucosal damage localized to the small intestine. Despite its prevalence, the only treatment currently available for celiac disease is complete avoidance of gluten proteins in the diet. Ongoing clinical trials have focused on targeting the immune response or gluten proteins through methods such as immunosuppression, enhanced protein degradation and protein sequestration. Recent studies suggest that polyphenols may elicit protective effects within the celiac disease milieu by disrupting the enzymatic hydrolysis of gluten proteins, sequestering gluten proteins from recognition by critical receptors in pathogenesis and exerting anti-inflammatory effects on the system as a whole. This review highlights mechanisms by which polyphenols can protect against celiac disease, takes a critical look at recent works and outlines future applications for this potential treatment method.

Impact of Atomizer Age and Flavor on In Vitro Toxicity of Aerosols from a Third-Generation Electronic Cigarette against Human Oral Cells.

Electronic cigarettes (ECs) are categorized into generations which differ in terms of design, aerosol production, and customizability. Current and former smokers prefer third-generation devices that satisfy tobacco cravings more effectively than older generations. Recent studies indicate that EC aerosols from first- and second-generation devices contain reactive carbonyls and free radicals and can cause in vitro cytotoxicity. Third-generation ECs have not been adequately studied. Further, previous studies have focused on cells from the respiratory tract, whereas those of the oral cavity, which is exposed to high levels of EC aerosols, have been understudied. We quantified the production of reactive carbonyls and free radicals by a third-generation EC and investigated the induction of cytotoxicity and oxidative stress in normal and cancerous human oral cell lines using a panel of eight commercial EC liquids. We found that EC aerosols produced using a new atomizer contained formaldehyde, acetaldehyde, and acrolein, but did not contain detectable levels of free radicals. We found that EC aerosols generated from only one of the eight liquids tested using a new atomizer induced cytotoxicity against two human oral cells in vitro. Treatment of oral cells with the cytotoxic EC aerosol caused a concomitant increase in intracellular oxidative stress. As atomizer age increased with repeated use of the same atomizer, carbonyl production, radical emissions, and cytotoxicity increased. Overall, our results suggest that third-generation ECs may cause adverse effects in the oral cavity and normal EC use, which involves repeated use of the same atomizer to generate aerosol, may enhance the potential toxic effects of third-generation ECs.

Loss and formation of malodorous volatile sulfhydryl compounds during wine storage.

Volatile sulfur compounds (VSCs), particularly low molecular weight sulfhydryls like hydrogen sulfide (H2S) and methanethiol (MeSH), are often observed in wines with sulfurous off-aromas. Recent work has shown both H2S and MeSH can increase up to a few µM (> 40 µg/L) during anoxic storage, but the identity of the latent sources of these sulfhydryls is still disputed. This review critically evaluates the latent precursors and pathways likely to be responsible for the loss and formation of these sulfhydryls during wine storage based on the existing enology literature as well as studies from food chemistry, geochemistry, biochemistry, and synthetic chemistry. We propose that three precursor classes have sufficient concentration and metastability to serve as latent sulfhydryl precursors in wine: 1) transition metal-sulfhydryl complexes, particularly those formed following Cu(II) addition, which are released under anoxic conditions through an unknown mechanism; 2) asymmetric disulfides, polysulfanes, and (di)organopolysulfanes formed through transition-metal mediated oxidation (e.g., Cu(II)) of sulfhydryls or pesticide degradation, and released through sulfitolysis, metal-catalyzed thiol-disulfide exchange or related reactions; 3) S-alkylthioacetates, primarily formed during fermentation, and releasable hydrolytically. Some evidence also exists for S-amino acids serving as precursors. Based on these findings, we propose a "decision tree" approach to choosing appropriate strategies for managing wines with sulfurous off-aromas.

Impact of electronic cigarette heating coil resistance on the production of reactive carbonyls, reactive oxygen species and induction of cytotoxicity in human lung cancer cells in vitro.

Electronic cigarette (e-cigarette; e-cig) use has grown exponentially in recent years despite their unknown health effects. E-cig aerosols are now known to contain hazardous chemical compounds, including carbonyls and reactive oxygen species (ROS), and these compounds are directly inhaled by consumers during e-cig use. Both carbonyls and ROS are formed when the liquid comes into contact with a heating element that is housed within an e-cig's atomizer. In the present study, the effect of coil resistance (1.5â€¯Ω and 0.25â€¯Ω coils, to obtain a total wattage of 8 ±â€¯2 W and 40 ±â€¯5 W, respectively) on the generation of carbonyls (formaldehyde, acetaldehyde, acrolein) and ROS was investigated. The effect of the aerosols generated by different coils on the viability of H1299 human lung carcinoma cells was also evaluated. Our results show a significant (p < 0.05) correlation between the low resistance coils and the generation of higher concentrations of the selected carbonyls and ROS in e-cig aerosols. Moreover, exposure to e-cig vapor reduced the viability of H1299 cells by up to 45.8%, and this effect was inversely related to coil resistance. Although further studies are needed to better elucidate the potential toxicity of e-cig emissions, our results suggest that these devices may expose users to hazardous compounds which, in turn, may promote chronic respiratory diseases.

Effects of Charcoal on Carbonyl Delivery from Commercial, Research, and Make-Your-Own Cigarettes.

Previous literature has shown that adding charcoal to cigarette filters can have varying effects on the delivery of toxic carbonyls depending on filter design, amount of charcoal, and puffing profiles. However, these studies have relied on either comparisons between commercially available charcoal and noncharcoal filtered cigarettes or experimental modification of filters to insert a charcoal plug into existing cellulose acetate filters. Make-your-own (MYO) cigarettes can help obviate many of the potential pitfalls of previous studies; thus, we conducted studies using commercial charcoal cigarettes and MYO cigarettes to determine the effects of charcoal on carbonyl delivery. To do this, we analyzed carbonyls in mainstream smoke by HPLC-UV after derivatization with 2,4-dinitrophenylhydrazine (DNPH). Charcoal was added in-line after the cigarettes or through the use of MYO charcoal cigarette tubes. MYO cigarettes had carbonyl deliveries similar to that of 3R4F research cigarette, regardless of tobacco type. The greatest effect on carbonyl delivery was observed with 200 mg of charcoal, significantly reducing all carbonyls under both methods tested. However, "on-tow" design charcoal filters, available on many commercially available charcoal brands, appeared to have a minimal effect on carbonyl delivery under intense smoking methods. Overall, we found that charcoal, when added in sufficient quantity (200 mg) as a plug, can substantially reduce carbonyl delivery for both MYO and conventional cigarettes. As carbonyls are related to negative health outcomes, such reductions may be associated with reductions in carbonyl-related harm in smokers.

Effects of Solvent and Temperature on Free Radical Formation in Electronic Cigarette Aerosols.

The ever-evolving market of electronic cigarettes (e-cigarettes) presents a challenge for analyzing and characterizing the harmful products they can produce. Earlier we reported that e-cigarette aerosols can deliver high levels of reactive free radicals; however, there are few data characterizing the production of these potentially harmful oxidants. Thus, we have performed a detailed analysis of the different parameters affecting the production of free radical by e-cigarettes. Using a temperature-controlled e-cigarette device and a novel mechanism for reliably simulating e-cigarette usage conditions, including coil activation and puff flow, we analyzed the effects of temperature, wattage, and e-liquid solvent composition of propylene glycol (PG) and glycerol (GLY) on radical production. Free radicals in e-cigarette aerosols were spin-trapped and analyzed using electron paramagnetic resonance. Free radical production increased in a temperature-dependent manner, showing a nearly 2-fold increase between 100 and 300 °C under constant-temperature conditions. Free radical production under constant wattage showed an even greater increase when going from 10 to 50 W due, in part, to higher coil temperatures compared to constant-temperature conditions. The e-liquid PG content also heavily influenced free radical production, showing a nearly 3-fold increase upon comparison of ratios of 0:100 (PG:GLY) and 100:0 (PG:GLY). Increases in PG content were also associated with increases in aerosol-induced oxidation of biologically relevant lipids. These results demonstrate that the production of reactive free radicals in e-cigarette aerosols is highly solvent dependent and increases with an increase in temperature. Radical production was somewhat dependent on aerosol production at higher temperatures; however, disproportionately high levels of free radicals were observed at ≥100 °C despite limited aerosol production. Overall, these findings suggest that e-cigarettes can be designed to minimize exposure to these potentially harmful products.

Influence of Smoking Puff Parameters and Tobacco Varieties on Free Radicals Yields in Cigarette Mainstream Smoke.

Cigarette smoke is a major exogenous source of free radicals, and the resulting oxidative stress is one of the major causes of smoking-caused diseases. Yet, many of the factors that impact free radical delivery from cigarettes remain unclear. In this study, we machine-smoked cigarettes and measured the levels of gas- and particulate-phase radicals by electron paramagnetic resonance (EPR) spectroscopy using standardized smoking regimens (International Organization of Standardization (ISO) and Canadian Intense (CI)), puffing parameters, and tobacco blends. Radical delivery per cigarette was significantly greater in both gas (4-fold) and particulate (6-fold) phases when cigarettes were smoked under the CI protocol compared to the ISO protocol. Total puff volume per cigarette was the major factor with radical production being proportional to total volume, regardless of whether volume differences were achieved by changes in individual puff volume or puff frequency. Changing puff shape (bell vs sharp vs square) or puff duration (1-5 s), without changing volume, had no effect on radical yields. Tobacco variety did have a significant impact on free radical production, with gas-phase radicals highest in reconstituted > burley > oriental > bright tobacco and particulate-phase radicals highest in burley > bright > oriental > reconstituted tobacco. Our findings show that modifiable cigarette design features and measurable user smoking behaviors are key factors determining free radical exposure in smokers.

Effect of Charcoal in Cigarette Filters on Free Radicals in Mainstream Smoke.

The addition of charcoal in cigarette filters may be an effective means of reducing many toxicants from tobacco smoke. Free radicals are a highly reactive class of oxidants abundant in cigarette smoke, and here we evaluated the effectiveness of charcoal to reduce free radical delivery by comparing radical yields from commercially available cigarettes with charcoal-infused filters to those without and by examining the effects of incorporating charcoal into conventional cigarette filters on radical production. Commercial cigarettes containing charcoal filters produced 40% fewer gas-phase radicals than did regular cellulose acetate filter cigarettes when smoked using the International Organization of Standardization (ISO, p = 0.07) and Canadian Intense (CI, p < 0.01) smoking protocols. While mean-particulate-phase radicals were 25-27% lower in charcoal cigarettes, differences from noncharcoal products were not significant ( p = 0.06-0.22). When cellulose acetate cigarette filters were modified to incorporate different types and amounts of activated charcoal, reductions in gas-phase (>70%), but not particulate-phase, radicals were observed. The reductions in gas-phase radicals were similar for the three types of charcoal. Decreases in radical production were dose-responsive with increasing amounts of charcoal (25-300 mg) with as little as 25 mg of activated charcoal reducing gas-phase radicals by 41%. In all studies, charcoal had less of an effect on nicotine delivery, which was decreased 33% at the maximal amount of charcoal tested (300 mg). Overall, these results support the potential consideration of charcoal in cigarette filters as a means to reduce exposure to toxic free radicals from cigarettes and other combustible tobacco products.

Little Cigars, Filtered Cigars, and their Carbonyl Delivery Relative to Cigarettes.

Introduction: Little cigars and filtered cigars are currently growing in popularity due to their low cost and wide variety of flavors while retaining an appearance similar to cigarettes. Given the health consequences associated with cigarette use, it is important to understand the potential harm associated with these similar products. This includes the potential harm associated with carbonyls (eg, acetaldehyde, acrolein, formaldehyde, etc.), an important class of toxicants and carcinogens in tobacco smoke. Our objective was to determine the carbonyl levels in mainstream smoke from little and filtered cigars compared to cigarettes. Methods: We examined two brands each of little cigars and filtered cigars, as well as two research cigarettes for carbonyl delivery using the International Organization of Standards (ISO) and the Health Canada Intense (HCI) machine-smoking protocols. Results: On a per puff basis, the levels of five of the seven carbonyls were higher from little cigars than filtered cigars and cigarettes (ISO: 56-116%; HCI: 39-85%; p < .05). On a per unit basis, most carbonyl levels were higher from both cigar types than cigarettes using the ISO method (ISO: 51-313%; p < .05) whereas only filtered cigars were higher using the HCI method (HCI: 53-99%; p < .05). Conclusion: These findings suggest that cigar smokers can be exposed to higher levels of carbonyls per cigar than cigarette smokers per cigarette. Implications: These data will increase our understanding of the relative harm from carbonyl exposure from little and filtered cigars both for cigar-only smokers and the cumulative harm among the growing population of cigarette-cigar multi-product smokers.

Effects of Topography-Related Puff Parameters on Carbonyl Delivery in Mainstream Cigarette Smoke.

Smoking topography parameters differ substantially between individual smokers and may lead to significant variation in tobacco smoke exposure and risk for tobacco-caused diseases. However, to date, little is known regarding the impact of individual puff parameters on the delivery of many harmful smoke constituents including carbonyls. To examine this, we determined the effect of altering individual puff parameters on mainstream smoke carbonyl levels in machine-smoked reference cigarettes. Carbonyls including formaldehyde, acetaldehyde, crotonaldehyde, propionaldehyde, methyl ethyl ketone (MEK), acrolein, and acetone were determined in cigarette smoke by HPLC after derivatization with 2,4-dinitrophenylhydrazine (DNPH). Deliveries of all carbonyls were nearly two-fold greater when cigarettes were smoked according to the more intense Health Canada Intense (HCI) protocol compared to the International Organization of Standardization (ISO) method, consistent with the two-fold difference in total puff volume between methods (ISO: 280-315 mL; CI: 495-605 mL). When individual topography parameters were assessed, changes in puff volume alone had the greatest effect on carbonyl delivery as predicted with total carbonyls being strongly correlated with overall puff volume (r2: 0.52-0.99) regardless of how the differences in volume were achieved. All seven of the carbonyls examined showed a similar relationship with puff volume. Minor effects on carbonyl levels were observed from vent blocking and changing the interpuff interval, while effects of changing puff duration and peak flow rate were minimal. Overall, these results highlight the importance of considering topography, especially puff volume, when the toxicant delivery and potential exposure smokers receive are assessed. The lack of an impact of other behaviors, including puff intensity and duration independent of volume, indicate that factors such as temperature and peak flow rate may have minimal overall effects on carbonyl production and delivery.

Variation in Free Radical Yields from U.S. Marketed Cigarettes.

Free radicals in tobacco smoke are thought to be an important cause of smoking-induced diseases, yet the variation in free radical exposure to smokers from different brands of commercially available cigarettes is unknown. We measured the levels of highly reactive gas-phase and stable particulate-phase radicals in mainstream cigarette smoke by electron paramagnetic resonance (EPR) spectroscopy with and without the spin-trapping agent phenyl-N-tert-butylnitrone (PBN), respectively, in 27 popular US cigarettes and the 3R4F research cigarette, machine-smoked according to the FTC protocol. We find a 12-fold variation in the levels of gas-phase radicals (1.2 to 14 nmol per cigarette) and a 2-fold variation in the amounts of particulate-phase radicals (44 to 96 pmol per cigarette) across the range of cigarette brands. Gas and particulate-phase radicals were highly correlated across brands (ρ = 0.62, p < 0.001). Both radicals were correlated with TPM (gas-phase: ρ = 0.38, p = 0.04; particulate-phase: ρ = 0.44, p = 0.02) and ventilation (gas- and tar-phase: ρ = -0.58, p = 0.001), with ventilation explaining nearly 30% of the variation in radical levels across brands. Overall, our findings of significant brand variation in free radical delivery under standardized machine-smoked conditions suggest that the use of certain brands of cigarettes may be associated with greater levels of oxidative stress in smokers.

Grape compounds suppress colon cancer stem cells in vitro and in a rodent model of colon carcinogenesis.

BACKGROUND: We have previously shown that the grape bioactive compound resveratrol (RSV) potentiates grape seed extract (GSE)-induced colon cancer cell apoptosis at physiologically relevant concentrations. However, RSV-GSE combination efficacy against colon cancer stem cells (CSCs), which play a key role in chemotherapy and radiation resistance, is not known. METHODS: We tested the anti-cancer efficacy of the RSV-GSE against colon CSCs using isolated human colon CSCs in vitro and an azoxymethane-induced mouse model of colon carcinogenesis in vivo. RESULTS: RSV-GSE suppressed tumor incidence similar to sulindac, without any gastrointestinal toxicity. Additionally, RSV-GSE treatment reduced the number of crypts containing cells with nuclear ß-catenin (an indicator of colon CSCs) via induction of apoptosis. In vitro, RSV-GSE suppressed - proliferation, sphere formation, nuclear translocation of ß-catenin (a critical regulator of CSC proliferation) similar to sulindac in isolated human colon CSCs. RSV-GSE, but not sulindac, suppressed downstream protein levels of Wnt/ß-catenin pathway, c-Myc and cyclin D1. RSV-GSE also induced mitochondrial-mediated apoptosis in colon CSCs characterized by elevated p53, Bax/Bcl-2 ratio and cleaved PARP. Furthermore, shRNA-mediated knockdown of p53, a tumor suppressor gene, in colon CSCs did not alter efficacy of RSV-GSE. CONCLUSION: The suppression of Wnt/ß-catenin signaling and elevated mitochondrial-mediated apoptosis in colon CSCs support potential clinical testing/application of grape bioactives for colon cancer prevention and/or therapy.

Enhanced phenylpyruvic acid production with Proteus vulgaris in fed-batch and continuous fermentation.

Phenylpyruvic acid is a deaminated form of phenylalanine and is used in various areas such as development of cheese and wine flavors, diagnosis of phenylketonuria, and to decrease excessive nitrogen accumulation in the manure of farm animals. However, reported phenylpyruvic acid fermentation studies in the literature have been usually performed at shake-flask scale with low production. In this study, phenylpyruvic acid production was evaluated in bench-top bioreactors by conducting fed-batch and continuous fermentation for the first time. As a result, maximum phenylpyruvic acid concentrations increased from 1350 mg/L (batch fermentation) to 2958 mg/L utilizing fed-batch fermentation. Furthermore, phenylpyruvic acid productivity was increased from 48 mg/L/hr (batch fermentation) to 104 and 259 mg/L/hr by conducting fed-batch and continuous fermentation, respectively. Overall, this study demonstrated that fed-batch and continuous fermentation significantly improved phenylpyruvic acid production in bench-scale bioreactor production.

Highly reactive free radicals in electronic cigarette aerosols.

Electronic cigarette (EC) usage has increased exponentially, but limited data are available on its potential harmful effects. We tested for the presence of reactive, short-lived free radicals in EC aerosols by electron paramagnetic resonance spectroscopy (EPR) using the spin-trap phenyl-N-tert-butylnitrone (PBN). Radicals were detected in aerosols from all ECs and eliquids tested (2.5 × 10(13) to 10.3 × 10(13) radicals per puff at 3.3 V) and from eliquid solvents propylene glycol and glycerol and from "dry puffing". These results demonstrate, for the first time, the production of highly oxidizing free radicals from ECs which may present a potential toxicological risk to EC users.

Screening of phenylpyruvic acid producers and optimization of culture conditions in bench scale bioreactors.

Alpha keto acids are deaminated forms of amino acids that have received significant attention as feed and food additives in the agriculture and medical industries. To date, their production has been commonly performed at shake-flask scale with low product concentrations. In this study, production of phenylpyruvic acid (PPA), which is the alpha keto acid of phenylalanine was investigated. First, various microorganisms were screened to select the most efficient producer. Thereafter, growth parameters (temperature, pH, and aeration) were optimized in bench scale bioreactors to maximize both PPA and biomass concentration in bench scale bioreactors, using response surface methodology. Among the four different microorganisms evaluated, Proteus vulgaris was the most productive strain for PPA production. Optimum temperature, pH, and aeration conditions were determined as 34.5 °C, 5.12, and 0.5 vvm for PPA production, whereas 36.9 °C, pH 6.87, and 0.96 vvm for the biomass production. Under these optimum conditions, PPA concentration was enhanced to 1,054 mg/L, which was almost three times higher than shake-flask fermentation concentrations. Moreover, P. vulgaris biomass was produced at 3.25 g/L under optimum conditions. Overall, this study demonstrated that optimization of growth parameters improved PPA production in 1-L working volume bench-scale bioreactors compared to previous studies in the literature and was a first step to scale up the production to industrial production.

Impact of pre- and post-fermentation fining with polyvinylpolypyrrolidone on the chemical stability and aromatic profile of Viognier wine.

Polyvinylpolypyrrolidone (PVPP) is a synthetic, insoluble polymer that can be added to white wines to improve the chemical stability of the final product by precipitating unstable low molecular weight phenolic compounds responsible for visual defects and undesirable flavor characteristics (e.g., excessive bitterness and/or astringency). The objective of this study was to characterize the effects of PVPP on the quality characteristics of Viognier wine when added pre- or post-fermentation as compared to an untreated control wine. Both PVPP-treated wines contained significantly lower concentrations of monomeric phenolics and browning pigments than the control wine (p ≤ 0.05). The addition of PVPP prior to fermentation conferred protection against oxidation of the wine as measured by acetaldehyde concentration (p ≤ 0.05). Analysis of the volatile aroma profile of each wine by headspace solid phase microextraction gas chromatographymass spectrometry (HS-SPME-GC-MS) revealed that the overarching aroma profiles of the PVPP-treated wines were significantly different from the control wine, but there was no difference between wines treated with PVPP pre-fermentation versus those treated post-fermentation. Specifically, statistically significant differences were observed in 9 of the 22 quantified aroma compounds, including those notably associated with the "stone fruit" aroma of Viognier. A negative correlation was identified between aroma compound concentration removal and the hydrophobicity of each compound, suggesting that the observed differences in aroma may be due to adsorption of aroma compounds by PVPP. The findings from this study present risks and benefits to wine quality upon treatment with PVPP at commercially recommended levels, and provide potentially valuable information for industrial wine producers.

Impact of mashing protocol on the formation of fermentable sugars from millet in gluten-free brewing.

The production of fermentable sugars (FS) in gluten-free (GF) brewing is hindered by the high starch gelatinization temperatures of GF malts and lower diastatic power compared to barley malt. Our previous work has demonstrated that starch gelatinization was the primary hurdle, and when decoupled from a single mash phase, high concentrations of FS could be produced. However, more research was required to improve the applicability of GF brewing. In this study, millet was used as a model GF malt demonstrating that despite the low α-amylase and ß-amylase activities compared to barley malt âˆ¼ 90 % of the FS (∼110 g/L) could be produced within 40 min. Limitations to enzyme extraction and separation due to coarse milling and lautering initially limited FS by âˆ¼ 30 g/L, requiring additional processing or exogenous enzyme supplements that improved fermentable sugar generation by âˆ¼ 20 g/L. Overall, millet is a promising brewing ingredient, provided appropriate mashing procedures are implemented.

Microscopic assessment of the degradation of millet starch granules by endogenous and exogenous enzymes during mashing.

The high gelatinization temperature (GT) of millet starch prevents the usage of infusion or step mashes as an effective means to generate fermentable sugars (FS) in brewing because the malt amylases lack thermostability at GT. Here, we investigate processing modifications to determine if millet starch can be efficiently degraded below GT. We determined that producing finer grists through milling did not introduce enough granule damage to markedly change gelatinization characteristics, though there was improved liberation of the endogenous enzymes. Alternatively, exogenous enzyme preparations were added to investigate their ability to degrade intact granules. At the recommended dosages (0.625 µL/g malt), significant FS concentrations were observed, although at lower concentrations and with a much-altered profile than possible with a typical wort. When exogenous enzymes were introduced at high (10×) addition rates, significant losses of granule birefringence and granule hollowing were observed well below GT, suggesting these exogenous enzymes can be utilized to digest millet malt starch below GT. The exogenous maltogenic α-amylase appears to drive the loss of birefringence, but more research is needed to understand the observed predominate glucose production.