Acetaldehyde Assay Kit

Many of the enzymatic test kits are official methods of prestigious organisations such as the Association of Official Analytical Chemicals (AOAC) and the American Association of Cereal Chemists (AACC) in response to the interest from oenologists. Megazyme decided to use its long history of enzymatic bio-analysis to make a significant contribution to the wine industry, by the development of a range of advanced enzymatic test kits. This task has now been successfully completed through the strategic and comprehensive process of identifying limitations of existing enzymatic bio-analysis test kits where they occurred, and then using advanced techniques, such as molecular biology (photo 1), to rapidly overcome them. Novel test kits have also been developed for analytes of emerging interest to the oenologist, such as yeast available nitrogen (YAN; see pages 2-3 of issue 117 article), or where previously enzymes were simply either not available, or were too expensive to employ, such as for D-mannitol analysis.

It is without doubt that testing plays a pivotal role throughout the whole of the vinification process. To produce the best possible quality wine and to minimise process problems such as “stuck” fermentation or troublesome infections, it is now recognised that if possible testing should begin prior to harvesting of the grapes and continue through to bottling. Traditional methods of wine analysis are often expensive, time consuming, require either elaborate equipment or specialist expertise and frequently lack accuracy. However, enzymatic bio-analysis enables the accurate measurement of the vast majority of analytes of interest to the wine maker, using just one piece of apparatus, the spectrophotometer (see previous issue No. 116 for a detailed technical review). Grape juice and wine are amenable to enzymatic testing as being liquids they are homogenous, easy to manipulate, and can generally be analysed without any sample preparation.

This study aimed to investigate the effect of mixture of herbal extracts and supplementary formula (FNP-C) on hangovers and antioxidant enzymes in alcohol-induced liver damage in rats. HepG2 cells were used as the experimental cells and divided into five groups: non-treated control (normal), alcohol-induced control (control), mixture of herbal extracts (FNP-B), FNP-C, and a commercial treatment of liver diseases (Livers^®); inhibition of detoxification and alcohol-induced damage was confirmed in vivo. Blood alcohol and acetaldehyde concentration after alcohol consumption were measured in a timely manner; alcohol dehydrogenase (ADH), aldehyde dehydrogenase (ALDH), superoxide dismutase (SOD), glutathione (GSH), glutathione transferase (GST), and lactate dehydrogenase (LDH) levels were measured in the liver. FNP-C exhibited the highest effect. When FNP-C was administered to alcohol-induced animals, blood alcohol and acetaldehyde concentration decreased compared to FNP-B and Livers^®. FNP-C reduced ADH levels and improved LDH, GSH, GST, and SOD levels. The FNP-C group was effective in preventing alcohol-induced hangovers and liver damage. Thus, FNP-C improves hangovers and increases antioxidant activity in an alcohol-induced model. Adding amino acids and vitamins to natural ingredients can potentially enhance the effect of improving hangovers.

The purpose of this study is to evaluate the effect of the bioconversion products of Oenanthe javanica extract fermented by Lactiplantibacillus plantarum (OEFL) on relieving hangovers and improving liver function. In addition, the bioactive substance of the OEFL, which alleviates hangover and ethanol-induced liver damage, was identified and its bioactive property was verified through in vivo experiments. In major substances analysis using high-performance liquid chromatography, OEFL produced 9.5-fold higher p-coumaric acid than the O. Javanica extract (OE). In addition, considering that quinic acid, which is not present in the OE, was produced in the OEFL it was confirmed that chlorogenic acid was decomposed into quinic acid by bioconversion. In the in vivo experiment using Sprague-Dawley rats, the OEFL and p-coumaric acid diets reduced blood ethanol, acetaldehyde, GPT, and ALP concentrations, increasing blood albumin concentrations compared to ethanol-administered groups, demonstrating that OEFL and p-coumaric acid, the main substance in the OEFL, improved ethanol-induced liver damage. Furthermore, the OEFL and its main bioactive substance, p-coumaric acid, alleviated liver fibrosis by downregulating TGF-β, SMAD-2, SMAD-4, α-SMA, and upregulating MMP-1. Therefore, OEFL is expected to be used as a functional food or pharmaceutical material as it has been confirmed to effectively relieve hangovers, prevent liver damage, and delay liver fibrosis in ethanol-induced liver damages.

Microoxygenation (MOX) is used to improve wine colour and sensory quality; however, limited information is available for Pinot noir wines and wines with different initial phenolic content. In this study, MOX was applied to two Pinot noir wines, with either a low or a high phenolic content, at two doses (0.50 and 2.11 mg/L/day) for 14 days. With the sterile filtration applied, acetaldehyde formation during MOX was very low, supporting the influence of yeast on acetaldehyde production during MOX. The MOX dosage rate did not significantly affect colour development, while the Pinot noir wine with higher phenolics benefited more from MOX, significantly increasing colour intensity and SO₂ resistant (polymeric) pigments. However, these changes did not guarantee colour stability, as a final SO₂ addition (100 mg/L) largely erased the improvement to colour in all wines. This could be due to the lower acetaldehyde formation, thus less ethyl-bridged stable pigments resistant to SO₂ bleaching. MOX also decreased the flavan-3-ols and anthocyanin monomers, which differed between the two Pinot noir wines, reflecting the initial phenolic content. Lastly, MOX generally increased the measured tannin concentration and affected the proportion of tannin subunits, with a decrease in tannin mass conversion and proportion of (-)-epigallocatechin extension units. Some of these changes in phenolic compounds could potentially increase astringency, suggesting that MOX should be applied to Pinot noir and other low phenolic wines with caution.

Zymomonas mobilis is an alpha-proteobacterium with a rapid ethanologenic pathway, involving Entner-Doudoroff (E-D) glycolysis, pyruvate decarboxylase (Pdc) and two alcohol dehydrogenase (ADH) isoenzymes. Pyruvate is the end-product of the E-D pathway and the substrate for Pdc. Construction and study of Pdc-deficient strains is of key importance for Z. mobilis metabolic engineering, because the pyruvate node represents the central branching point, most novel pathways divert from ethanol synthesis. In the present work, we examined the aerobic metabolism of a strain with partly inactivated Pdc.

This study investigates the influence of the antimicrobial agent chitosan on a selected Schizosaccharomyces pombe strain during the alcoholic fermentation of ultra-pasteurized grape juice with a high concentration of malic acid. It also studies a selected Saccharomyces cerevisiae strain as a control. The study examines several parameters relating to wine quality, including volatile and non-volatile compounds. The principal aim of the study is to test the influence of chitosan on the final chemical composition of the wine during alcoholic fermentation, and to compare the two studied fermentative yeasts between them. The results show that chitosan influences the final concentration of acetic acid, ethanol, glycerol, acetaldehyde, pyruvic acid, α-ketoglutarate, higher alcohols, acetate esters, ethyl esters, and fatty acids, depending on the yeast species.

China has a large market for dried persimmon fruits, but mold and sulfur dioxide (SO₂) residue have become serious potential consumer health risks. To reduce these risks, the adopted ozone and/or UV-C sterilization methods were applied; however, these methods had low effectiveness due to lack of sterilization condition optimization based on the drying process. In this study, bacteria and fungi strains were identified and optimal ozone and/or UV-C conditions were selected by treating predominant strains. Results show that Curtobacterium, Luteibacter, Rhodotorula, Penicillium, and Aspergillus were the predominant microorganisms in the whole artificial drying process, and drying at 36 and 72 h were found to effectively influence the quantity of microorganism contamination. Comparing the mortalities of microorganism revealed that the sterilization of combining ozone with UV-C was superior to the individual utilization of ozone or UV-C. Moreover, the physic-chemical results of the dried persimmon fruits suggested that this non-thermal sterilization could alleviate astringency, but hadn't significant effects on other properties, including color, moisture content, water activity, and protopectin. Therefore, these optimal conditions could potentially enhance the production of dried persimmon fruits.

This study is purposed to check up the natural 12 kinds of herbal extracts suitable for hangover and based on the results of contents of phenolic compounds, ABTS radical scavenging activity, alcohol dehydrogenase (ADH), and acetaldehyde dehydrogenase (ALDH). Selected 8 kinds of herbal extracts are blended according to the efficacy and the pearson's correlation between each content and activity. C. sinensis var. sinensis, P. densiflora Gnarl and P. lobata Ohwi showed excellent ADH activity, P. lobata Ohwi had a strong correlation between the content and efficacy, and C. sinensis var. sinensis, P. densiflora Gnarl had a negative correlation. Through the ADH and ALDH activity test of F.1 to F.7, the F.7 showed the highest synergic effect and selected as an optimal formulation. F.7 intake-group, the breath alcohol concentration was significantly reduced to 58% after 30 minutes and 27% after 120 minutes, compared to right after alcohol consumption. After alcohol consumption, there was a significant improvement effect (p<0.05) in tired and thirst in the intake group compared to the non-intake group.

Hepatic protective effects of Ligularia fischeri (LF) and Aronia melanocarpa (AM) against alcohol were investigated in vitro and in vivo test. LF, AM, and those composed mixing material (LF+AM) were treated in HepG2 cell. Alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH) activities were significantly increased in each singleness extract and mixed composite. The protective effect on alcoholic liver damage was investigated by animal models. Serum alcohol level and acetaldehyde level were significantly decreased by LF+AM treatment in acute experimental model. In the chronic mouse model study, we had found that the increased plasma liver damage index (alkaline phosphatase) by alcohol treatment was declined by oral administration of LF+AM extraction composite. As well as, it was identified that the protection effect was induced by increasing catalase activity and suppressing COX-2, TNF-α, MCP-1, and IL-6 mRNA expressions. CYP2E1 mRNA expression was also increased. These results suggest that oral ingestion of LF and AM mixed composite is able to protect liver against alcohol-induced injury by increasing alcohol metabolism activity and antioxidant system along with decreasing inflammatory responses.

Use of chronic alcohol produces alcoholic liver disease, with relationship of the beginning of abnormal lipid metabolism. Recent studies show that abnormal cholesterol metabolism has a specific role in the pathological cause of alcoholic fatty liver disease. Gymnaster koraiensis (GK), a worthy perennial Korean wild plant has specific polyacetylene compounds. Many researches have presented that GK has many pharmacological properties, such as oxidation prevention, liver protection, and inflammation prevention. However, the conservative effect of GK on alcoholic fatty liver has not been researched so far. Male Sprague-Dawley rats were randomly separated to normal feeding (fed a normal feeding for 4 weeks) and ethanol feeding (ED) groups. Rats in the ED group were administered a Lieber-DeCarli liquid feeding (containing 6.7% ethanol) and administered GK extract (125, 250, or 500 mg/kg/day), silymarin (200 mg/kg/day), or no treatment for 4 weeks. Each treatment group contained six rats. The administration with GK decreased serum levels of triglycerides, alcohol, alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase, while increased serum level of adiponectin and activity of alcohol dehydrogenase. In alcohol-triggered fatty liver, GK decreased total cholesterol and fatty acid synthase, while increased catalase and superoxide dismutase. Alterations in liver histology, as assessed by H&E staining, showed that the GK treatment decreased amass of lipids in liver. These results present that GK extract may be potential therapeutic agent for alcoholic fatty liver disease by preventing fatty acid synthesis and activating anti-oxidation enzymes, while in activating alcohol degradation in ethanol-triggered fatty liver.

Background: Human placenta extract (HPE) has been used to treat a number of liver diseases. Porcine placenta is relatively safe and has been reported to have similar immune effects to HPE and used as its alternative. This study evaluates the effect of enzymatic porcine placental extract (EPPE, Uni‐Placenta^®) on alcohol pharmacokinetics in rat. Methods: This study was designed to determine the effect of single‐dose EPPE on the pharmacokinetics of alcohol and liver function. Results were based on serum alcohol and acetaldehyde concentrations and activities of hepatic and gastric ADH and ALDH in rats. Results: The hepatic ADH in alcohol group was significantly increased and it may be enzyme‐induction by alcohol. The hepatic ALDH and gastric ADH were not changed, but gastric ALDH was significantly decreased only in the high‐dose EPPE group. In the alcohol pharmacokinetics parameters, the AUC was 44.5 mM∙h in the alcohol group. Otherwise, AUCs of low, middle, high, and silymarin groups were significantly decreased. C_max was reached at 1 hour and then gradually decreased to 63% and 43% in the middle and high groups at 3 hours, respectively, and to 92% in the low groups. The pharmacokinetics and serum concentrations of acetaldehyde showed no differences between EPPE groups except the silymarin group. No histologic changes were seen in any group. Conclusions: The single‐dose EPPE (0.5 to 2.5 g/kg) suppressed absorption of alcohol in the gastrointestinal tract. This may be useful in preventing hangover effects and toxicity after drinking alcohol and may also preserve liver health after alcohol ingestion.

Osteopenia or osteoporosis occurs frequently in alcoholics and patients with alcoholic fatty liver disease. Methoxsalen (MTS), 8-methoxypsoralen, improved osteoporosis in ovariectomized and diabetic mouse models; however, its effects on alcohol-induced osteopenia and steatosis have not been reported. This study examined the effects of MTS on alcohol-induced bone loss and steatosis. Rats in the alcohol groups were fed a Liber-DeCarli liquid diet containing 36% of its calories as alcohol. MTS was at 0.005% in their diet, while alendronate (positive control; 500 μg/kg BW/day) was administered orally for eight weeks. The pair-fed group received the same volume of isocaloric liquid diet containing dextrin-maltose instead of alcohol as the alcohol control group consumed the previous day. In the alcohol-fed rats, the MTS and alendronate increased the bone volume density, bone surface density and trabecular number, while the bone specific surface, trabecular separation and structure model index were decreased in the tibia. MTS down-regulated tibial tartrate-resistant acid phosphatase 5 (TRAP) expression compared to the alcohol control group. MTS or alendronate prevented chronic alcohol-induced hepatic lipid accumulation and the triglyceride level in the alcohol-fed rats by decreasing the lipogenic enzyme activities and increasing the fatty acid oxidation enzyme activities. MTS reduced significantly the serum levels of alcohol, TRAP and tumor necrosis factor-α compared to the alcohol control group. Overall, these results suggest that MTS is likely to be an alternative agent for alcoholic osteopenia and hepatosteatosis.

Liver X receptor (LXR) agonists have the potential to alleviate obesity related diseases, particularly atherosclerosis. However, LXRs are transcriptional regulators that induce de novo lipogenesis and lipid accumulation in hepatocytes which represents a serious adverse effect. In this work, we sought to characterize the LXR agonist GW3965 effects on fatty acid (FA) and phospholipid (PL) remodelling and the correlation with gene expression in order to better understand the underlying effects leading to hepatic pathology upon LXR activation. Human primary hepatocytes treated for 48 h with GW3965 were analysed for changes in lipid metabolism gene expression by qPCR, variations in the FA profile was evaluated by GC-FID and in PL profiles using thin layer chromatography, ESI-MS and MS/MS analysis. Changes in cell membrane biochemical properties were studied using bilayer models generated with CHARMM-GUI. ELOLV6 and SCD1 mRNA increase was consistent with higher C16:1 and C18:1n9 at the expense of C16:0 and C18:0. The reduction of C18:2n6 and increase in C20:2n6 was in agreement with ELOVL5 upregulation. Phosphatydilethanolamine (PE) levels tended to decrease and phosphatidylinositol to increase; although differences did not reach significance, they correlated with changes in AGXT2L1, CDS1 and LPIN1 mRNA levels that were increased. The overall effect of GW3965 on PEs molecular profiles was an increase of long-chain polyunsaturated FA chains and a decrease of C16/C18 saturated and monounsaturated FAs chains. Additionally, PC (32:1) and PC (34:2) were decreased, and PC (36:1) and PC (34:1) were increased. AGXT2L1 is an enzyme with strict substrate specificity for phosphoethanolamine, which is converted into ammonia in GW3965-treated hepatocytes and could explain the PE reduction. In summary, LXR activation by GW3965 targets PE biosynthesis and FA elongation/desaturation, which tends to decrease PE in relation to total PL levels, and remodelling of PC and PE molecular species. We identified the human AGXT2L1 gene as induced by LXR activation by both synthetic and endogenous agonist treatment. The increase in acetaldehyde-induced oxidative stress, and in the lipid species identified have the potential to enhance the inflammatory process and impair membrane function. Future studies should focus on inhibition of AGXT2L1 activity with the aim of reverting the steatosis induced by LXR activation.

Ecological niche difference associated with varied ethanol tolerance between Drosophila suzukii and Drosophila melanogaster (Diptera: Drosophilidae). Florida Entomologist, 101(3), 498-504.

Drosophila suzukii (Matsumura) (Diptera: Drosphilidae) is an important pest that causes damage to fruits of over 60 plant species. Drosophila suzukii oviposits on ripe fruit, while D. melanogaster oviposits on decaying fruit. Therefore, these species occupy separate ecological niches. To provide a better understanding of the alcohol tolerance between these 2 species and explore the relationship of ecological niche differences and alcohol tolerance, ethanol and acetaldehyde content was examined in red grapes infested by D. melanogaster and D. suzukii. We assessed mortality and alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH) activity levels for 2 Drosophila species exposed to ethanol. The study results showed that ethanol content gradually increased as the fruit decayed while being infested by Drosophila. The ethanol content was higher in the presence of D. melanogaster than in the presence of D. suzukii. In the mortality experiment, the LC₅₀ of D. melanogaster adults was approximately 8.0% following exposure to ethanol for more than 6 h, while it was only 2.7% in D. suzukii. Moreover, D. melanogaster adults and larvae all had higher ADH and ALDH activity than D. suzukii exposed to ethanol. Our results suggest that D. melanogaster and D. suzukii may occupy different ecological niches due to their discrepancy in tolerance to environmental ethanol, which is mainly regulated by ADH and ALDH.

The aim of this study was to investigate the effects of mixed inoculation on blocking astringency in aronia (Aronia melanocarpa) concentrates with respect to chemical and sensory properties. Mixed inoculation treatment included inoculation with four different ratios of mixed starter cultures (Lactobacillus bulgaricus, Streptococcus thermophilus, and Saccharomyces cerevisiae). The effect on astringency in aroina concentrates after mixed inoculation was demonstrated by three intimately reciprocal tests: bovine serum albumin precipitation, condensed tannin, and acetaldehyde analyses. After mixed inoculation, increased acetaldehyde components in aronia concentrates reacted with condensed tannin to form the modified tannin, resulting in decreased condensed tannin contents. This trend was supported by reduced aggregation of tannin-protein complex. Furthermore, sensory analyses (consumer testing and ranking descriptive analysis) showed moderated astringency perceptions after mixed inoculation. For consumer testing, the liking scores of all sensory attributes (overall liking, astringency, bitterness, sweetness, sourness, and berry taste) in mixed inoculated samples were slightly higher than those of non-inoculated aronia concentrate. The findings of ranking descriptive analysis indicated the significantly decreased scores at astringency attribute after mixed inoculation. In conclusion, these results suggested that mixed inoculation can be used to moderate astringency in aronia concentrates in both chemical and sensory aspects.

This study aims to investigate the protective effects of gastrodin (GSTD), a natural compound isolated from the root of Gastrodia elata BL., on ethanol-induced liver injury and apoptosis in HepG2 cells and animal models. For in vitro studies, GSTD was used to pre-treat the cells for 4 h followed by 600 mM of ethanol co-administration for 24 h. Alcoholic liver disease (ALD) of Sprague-Dawley (SD) rats was induced by chronic ethanol-feeding plus a single dose (5 g/kg) of acute ethanol administration, GSTD at different doses were co-administered for 8 weeks. For acute liver injury experiment of ICR mice, GSTD (100 mg/kg/d) was pre-treated for 3 d followed by ethanol administration (5 g/kg) for 3 times. The results showed that GSTD protects HepG2 cells from ethanol-induced toxicity, injury, and apoptosis significantly. Co-administered with ethanol, GSTD prevented the loss of mitochondrial membrane potential, reduced the release cytochrome c from mitochondria, and inhibited the activation of caspase-3 in HepG2 cells. In SD rats induced by chronic ethanol-feeding, GSTD significantly restored liver function and ameliorated pathological changes of the liver. In rat liver, GSTD greatly suppressed the activation of caspase-3 and inhibited hepatocellular apoptosis. In ethanol-induced acute liver injury of ICR mice, GSTD reduced liver acetaldehyde and suppressed the up-regulation of alcohol dehydrogenase (ADH) and CYP2E1 significantly. Our results demonstrate that GSTD is efficacious in protecting liver cells from ethanol-induced injury and apoptosis; it may be useful for the development of novel agents for the treatment of ALD in the future.

Two different Metschnikowia strains (M. pulcherrima MP 346 or M. fructicola MF 98-3) were applied for the first time, during pre-fermentative cold maceration (PCM) in order to enhance the properties and stability of Sangiovese wine color. During the 2014 and 2015 vintages a total of eight wines were produced with 24 h of cold maceration (PCM 24 h) or 72 h (PCM 72 h), respectively. PCM was carried out in presence of MP 346 or MF 98-3 or pectic enzyme (Cuvée Rouge). The sequential inoculation of S. cerevisiae strain was carried out at the end of PCM. After 12 months in the bottle, the MP 346 and MF 98-3 wines contained much higher levels of total flavonoids than the Control sample for both vintages and regardless PCM duration. Moreover, in both vintages only MF 98-3 showed a higher color intensity than the Control sample after 12 months in the bottle. However, neither PCM duration nor the microbial/enzymatic treatment increased the level of anthocyanins at draining off. Both wines produced by the pre-fermentative inoculum with Metschnikowia strains (MP 346 and MF 98-3) retained their red hue, regardless the duration of pre-fermentative and fermentative macerations, while the Control wines were characterized by faster rates of color loss.

Acetic acid is undesired in Icewine. It is unclear whether its production by fermenting yeast is linked to the nicotinamide adenine dinucleotide (NAD⁺/NADH) system or the nicotinamide adenine dinucleotide phosphate (NADP+/NADPH) system. To answer this question, the redox status of yeast cytosolic NAD(H) and NADP(H) were analyzed along with yeast metabolites to determine how redox status differs under Icewine versus table wine fermentation. Icewine juice and dilute Icewine juice were inoculated with commercial wine yeast Saccharomyces cerevisiae K1-V1116. Acetic acid was 14.3-fold higher in Icewine fermentation than the dilute juice condition. The ratio of NAD⁺ to total NAD(H) was 24-fold higher in cells in Icewine fermentation than the ratio from the dilute juice condition. Conversely, the ratio of NADP⁺ to total NADP(H) from the dilute fermentation was 2.9-fold higher than that in the Icewine condition. These results support the hypothesis that in Icewine, increased NAD⁺ triggered the catalysis of NAD⁺-dependent aldehyde dehydrogenase(s) (Aldp(s)), which led to the elevated level of acetic acid in Icewine, whereas, in the dilute condition, NADP⁺ triggered NADP⁺-dependent Aldp(s), resulting in a lower level of acetic acid. This work, for the first time, analyzed the yeast cytosolic redox status and its correlation to acetic acid production, providing a more comprehensive understanding of the mechanism of acetic acid production in Icewine.

Vinegar is an inexpensive commodity, and economic considerations require that a relatively low-cost raw material be used for its production. An investigation into the use of a new, alternative substrate - pineapple waste - is described. This approach enables the utilization of the pineapple’s (Ananas comosus) peels and core, which are usually discarded during the processing or consumption of the fruit. Using physical and enzymatic treatments, the waste was saccharified, and the resulting substrate was fermented with Saccharomyces cerevisiae for 7-10 days under aerobic conditions at 25°C. This resulted in an alcohol yield of approximately 7%. The alcoholic medium was then used as a seed broth for acetic fermentation using Acetobacter aceti as the inoculum for approximately 30 days at 32°C to obtain 5% acetic acid. Samples were analyzed at the beginning and end of the acetification cycle to assess the volatile and fixed compounds by GC-MS and UHPLC-QTOF-MS. The metabolomic analysis indicated that L-lysine, mellein, and gallic acid were significantly more concentrated in the pineapple vinegar than in the original wine. Higher alcohols, aldehydes, and ketones characterized the aroma of the final pineapple vinegar, whilst off-flavors were significantly reduced relative to the initial wine. This study is the first to highlight the metabolite profile of fruit vinegar with a slight floral aroma profile derived from pineapple waste. The potential to efficiently reduce the post-harvest losses of pineapple fruits by re-using them for products with added food values is also demonstrated.