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Detecting ethanol and acetaldehyde by simple and ultrasensitive fluorimetric methods in compound foods
Abstract
Abstract
There is a need for simple, accurate, and rapid analysis of ethanol (Eth) and acetaldehyde (AA) in a wide variety of beverages and foods. A novel enzymatic assay coupled to formation of fluorescent chromophore is presented. Eth detection was further improved by adding semicarbazide to the reaction mixture, which interacts with AA and prevents its inhibitory effect on Eth oxidation. The limits of detection of Eth (0.5 mg/L) and AA (0.9 mg/L) are comparable with the performance of modern gas chromatography techniques. The repeatability of Eth and AA detection in various foods (9% on average) was lower than that with commercial kits (23%). The high sensitivity of the developed method enables detection of AA in common foods [e.g., bio-yogurt (12.2 mg/L), and the existence of endogenous Eth (1.8 mg/L) and AA (2.0 mg/L) in bacteria-free non-fermented bovine milk], which could not measured so far by enzymatic methods.
Highlights
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A method for assaying ethanol and acetaldehyde in compound foods is described.
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Ethanol and acetaldehyde can be measured in wide varieties of beverages and foods.
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The detection limit allows finding of mutagenic level of acetaldehyde in samples.
... Ethanol is commonly present in disinfectant, beverage, refrigerant, and it becomes an important reagent in the production of biodiesel [1][2][3][4][5][6][7][8][9][10][11][12][13][14] . Declaration of ethanol content in the product's label is really required for the regulation and health risk purposes. ...
... Therefore, ethanol content monitoring and adsorption due to environmental issues are necessary for the quality control of the products. Ethanol quantification has been done using spectrofluorometer, Fourier transform infrared (FT-IR) spectroscopy, near-infrared (NIR) spectroscopy, and gas chromatography measurements [5][6][7] . Those methods have required large-space and high-technology apparatus which could not be directly used in portable applications. ...
By using gold(I) 4-(3,5-dimethoxybenzyl)-3,5-dimethyl pyrazolate complex as a chemosensor, we report a successful quantification of ethanol in aqueous solution based on vapor-induced phosphorescent quenching. Upon monitoring at 278 nm irradiation, the emission intensity at 609 nm of the complex was quenched by the presence of ethanol vapors from the mixture. It was found that the intensity is linearly decreased with increment of ethanol concentration where limit of detection was found to be 11.7% v/v in ethanol/water. When the chemosensor was applied to quantify ethanol content of 4 real samples, the accuracy was achieved up to 96%, which is remarkable.
... Similarly, various concentrations of ethanol were produced on the surface of food items such as milk, fruits, and vegetables due to microbial activities. 9,12,13 Thus, monitoring ethanol vapor from our breath and the surface of food items can be used for noninvasive diagnosis and freshness level of food items. In this scenario, the design and development of sensors with desired response characteristics with which to detect ethanol at an ambient atmosphere has a clear mandate. ...
The article's abstract is not available.
Background:
Alcohol is a risk factor for cancer of the oral cavity, pharynx, oesophagus, colorectum, liver, larynx and female breast, whereas its impact on other cancers remains controversial.
Methods:
We investigated the effect of alcohol on 23 cancer types through a meta-analytic approach. We used dose-response meta-regression models and investigated potential sources of heterogeneity.
Results:
A total of 572 studies, including 486 538 cancer cases, were identified. Relative risks (RRs) for heavy drinkers compared with nondrinkers and occasional drinkers were 5.13 for oral and pharyngeal cancer, 4.95 for oesophageal squamous cell carcinoma, 1.44 for colorectal, 2.65 for laryngeal and 1.61 for breast cancer; for those neoplasms there was a clear dose-risk relationship. Heavy drinkers also had a significantly higher risk of cancer of the stomach (RR 1.21), liver (2.07), gallbladder (2.64), pancreas (1.19) and lung (1.15). There was indication of a positive association between alcohol consumption and risk of melanoma and prostate cancer. Alcohol consumption and risk of Hodgkin's and Non-Hodgkin's lymphomas were inversely associated.
Conclusions:
Alcohol increases risk of cancer of oral cavity and pharynx, oesophagus, colorectum, liver, larynx and female breast. There is accumulating evidence that alcohol drinking is associated with some other cancers such as pancreas and prostate cancer and melanoma.
The assumption that metabolites derived from the activity of the mammary gland epithelial cells reflects changes in milk secretion and its coagulation properties was tested in dairy cows. The experiment included cows with uninfected udders and cows with one of the glands infected by different bacteria specie. Analysis were carried at the cow level (including all four glands), or at the gland level. High and significant correlations among the concentrations of lactose, glucose, glucose-6-posphate, milk related respiratory index (the ratio between the concentrations of citrate/lactate+malate in milk) and milk-derived glycolytic index (the ratio between glucose-6-phosphate and glucose in milk) and milk clotting parameters were found. The physiological basis for these relations and their ability to predict the deterioration in milk quality in subclinically infected glands and in glands previously clinically infected with Escherichia coli are discussed.
The effects of separate infection with four major pathogens frequently associated with the occurrence of subclinical mastitis in cows (Staphylococcus aureus, S. chromogenes, Escherichia coli and Streptococcus dysgalactiae) on milk quality for cheese production were studied for quarters of the same animal. Infection increased somatic cell count (SCC), modified leucocyte distribution, decreased lactose concentration and increased proteolysis of casein. Regardless of bacteria type, the plasmin activity in milk from the infected glands increased ∼2 fold compared with uninfected quarters. These changes were associated with increased rennet clotting time and decreased curd firmness for milk from infected glands, indicating that milk quality for cheese production was negatively affected by infection. Although the general pattern of bacterial invasion was similar, each type of bacteria elicited the above-described responses in a specific manner. SCC, commonly used by the dairy industry as a measure of milk hygienic quality, provided the poorest prediction of milk quality for cheese production in comparison to indices of proteolysis of casein.
Background:
Methanol, acetaldehyde, acetone, and ethanol, which are commonly used as biomarkers of several diseases, in acute intoxications, and forensic settings, can be detected and quantified in biological fluids. Gas chromatography (GC)-mass spectrometry techniques are complex, require highly trained personnel and expensive materials. Gas chromatographic determinations of ethanol, methanol, and acetone have been reported in one study with suboptimal accuracy. Our objective was to improve the assessment of these compounds in human blood using GC with flame ionization detection.
Methods:
An amount of 50 µl of blood was diluted with 300 µl of sterile water, 40 µl of 10% sodium tungstate, and 20 µl of 1% sulphuric acid. After centrifugation, 1 µl of the supernatant was injected into the gas chromatograph. We used a dimethylpolysiloxane capillary column of 30 m × 0.25 mm × 0.25 µm.
Results:
We observed linear correlations from 7.5 to 240 mg/l for methanol, acetaldehyde, and acetone and from 75 to 2400 mg/l for ethanol. Precision at concentrations 15, 60, and 120 mg/l for methanol, acetaldehyde, and acetone and 150, 600, and 1200 mg/ml for ethanol were 0.8-6.9%. Ranges of accuracy were 94.7-98.9% for methanol, 91.2-97.4% for acetaldehyde, 96.1-98.7% for acetone, and 105.5-111.6% for ethanol. Limits of detection were 0.80 mg/l for methanol, 0.61 mg/l for acetaldehyde, 0.58 mg/l for acetone, and 0.53 mg/l for ethanol.
Conclusion:
This method is suitable for routine clinical and forensic practices.
The occurrence and persistence of diacetyl in samples of unfermented raw and processed milk have been investigated; in addition, samples of milk fermented by different lactic acid bacteria have also been analysed. Diacetyl was determined by using a simplified gas chromatographic method in which acetone was the extracting agent. Diacetyl in unfermented raw milk of cow, buffalo, goat and sheep was found to be 45.1, 65.2, 42.8 and 41.2 mg kg−1, respectively. The pre-fermentative diacetyl in milk was highly stable, and its initial value irrelevantly changed after the milk had been stored at 20 °C for 24 h or freeze-dried for 8 h and finally, having been thermically treated at 80 °C for 30 min. Significant amounts of diacetyl were detected also in all the processed milks. At 26 °C growth temperature, the tested lactic acid bacteria produced diacetyl in milk in the following order: Streptococcus thermophilus < Lactobacillus paracasei < Lactobacillus rhamnosus. Diacetyl increased during fermentation of 24 h, while a spontaneous and significant loss was registered when fermentation was prolonged to 48 h.
Background: Esophageal cancer is unusually frequent in Western Kenya, despite the low prevalence of classical risk factors such as heavy drinking and tobacco smoking. Among Kenyans consumption of fermented milk is an old tradition. Our hypothesis is that alcohol and acetaldehyde are produced during the fermentation process and that their carcinogenic potential contributes to the high incidence of esophageal cancer.
Methods: Eight samples of mursik milk starter cultures were collected from different Kalenjin families in the Rift Valley province, Western Kenya. A protocol provided by the families was used for milk fermentation. Ethanol and acetaldehyde levels were measured by gas chromatography. The microbial flora in starter cultures was identified by 16S and 18S sequencing.
Results: 7/8 starter cultures produced mutagenic (>100 μmol/L) levels of acetaldehyde and 4/8 starter cultures produced more than 1,000 μmol/L of acetaldehyde. The highest alcohol levels (mean 79.4 mmol/L) were detected in the four fermented milks with highest acetaldehyde production. The mean number of microbial species in the starter cultures was 5 (range 2–8). Yeasts were identified in all starter cultures (mean 1.5 species/milk) but their proportion of the total microbial count varied markedly (mean 35%, range 7%–90%). A combination of yeast and lactobacilli, especially Candida krusei with Lactobacillus kefiri, with the exclusion of other species, seemed to correlate with higher acetaldehyde and ethanol levels.
Conclusions: Significant levels of ethanol and acetaldehyde were produced during mursik fermentation.
Impact: When ingested several times daily the repeated exposure to carcinogenic levels of acetaldehyde may contribute to esophageal carcinogenesis.
Alcohol consumption is one of the top 10 risks for the worldwide burden of disease and an established cause of head and neck cancer, as well as cancer at other sites. Acetaldehyde, the major metabolite of ethanol, reacts with DNA to produce adducts, which are critical in the carcinogenic process and can serve as biomarkers of exposure and, possibly, of disease risk. Acetaldehyde associated with alcohol consumption is considered "carcinogenic to humans." We have previously developed the technology to quantify acetaldehyde-DNA adducts in human tissues, but there are no studies in the literature defining the formation and removal of acetaldehyde-DNA adducts in people who consumed alcohol.
We investigated levels of N(2)-ethylidene-dGuo, the major DNA adduct of acetaldehyde, in DNA from human oral cells at several time points after consumption of increasing alcohol doses. Ten healthy nonsmokers were dosed once a week for three weeks. Mouthwash samples were collected before and at several time points after the dose. N(2)-Ethylidene-dGuo was measured as its NaBH(3)CN reduction product N(2)-ethyl-dGuo by liquid chromatography-electrospray-tandem mass spectrometry.
N(2)-ethylidene-dGuo levels increased as much as 100-fold from baseline within 4 hours after each dose for all subjects and in a dose-responsive manner (P = 0.001).
These results show an effect of alcohol on oral cell DNA adduct formation, strongly supporting the key role of acetaldehyde in head and neck cancer caused by alcohol drinking.
Our results provide some of the first conclusive evidence linking exposure to a lifestyle carcinogen and kinetics of DNA adduct formation in humans.
Support of milk production in modern dairy cows demands a large proportion of its own metabolic resources, such as glucose, which might be required under stressful situations. The aim of the experiment was to test the hypothesis that acute immune stress shifts oxidative metabolism to glycolysis. Two mammary quarters in 6 Holstein cows were infused with lipopolysaccharide (LPS), whereas the 2 counter quarters served as controls to the treatment. An additional 6 cows were infused with saline and served as running controls. The LPS challenge induced dramatic transient increases in milk lactate (75-fold) and malate (11-fold) concentrations (both markers of glycolysis) at 24h posttreatment. No significant changes in lactate and malate concentrations were recorded in control quarters and control animals, indicating that the effect of LPS was restricted to the treated gland. The LPS challenge induced a dramatic transient decrease in milk yield, and lactose and citrate (a marker of mitochondrial metabolism) secretion at 24h posttreatment. The kinetics were inversely proportional to those of lactate and malate concentrations. Thus, our data suggest that LPS challenge induces acute conversion of epithelial cell metabolism from principally mitochondrial-oxidative to principally cytosolic (glycolytic), which allows the diversion of metabolic resources normally used to synthesize milk to support the immune system. An in vitro bacterial growth test showed that concentrations of lactate, malate, and lactose equivalent to those found in the in vivo experiment delayed and reduced the growth of a pathogenic Escherichia coli strain, suggesting that they play a role in diminution of bacterial multiplication in the mammary gland.
Acetaldehyde (ethanal) is a genotoxic carcinogen, which may occur naturally or as an added flavour in foods. We have developed an efficient method to analyze the compound in a wide variety of food matrices. The analysis is conducted using headspace (HS) gas chromatography (GC) with flame ionization detector. Using a robot autosampler, the samples are digested in full automation with simulated gastric fluid (1 h at 37°C) under shaking, which frees acetaldehyde loosely bound to matrix compounds. Afterwards, an aliquot of the HS is injected into the GC system. Standard addition was applied for quantification to compensate for matrix effects. The precision of the method was sufficient (<3% coefficient of variation). The limit of detection was 0.01 mg/L and the limit of quantification was 0.04 mg/L. 140 authentic samples were analyzed. The acetaldehyde content in apples was 0.97 ± 0.80 mg/kg, orange juice contained 3.86 ± 2.88 mg/kg. The highest concentration was determined in a yoghurt (17 mg/kg). A first-exposure estimation resulted in a daily acetaldehyde intake of less than 0.1 mg/kg bodyweight from food, which is considerably lower than the exposures from alcohol consumption or tobacco smoking.
An increasing body of evidence now implicates acetaldehyde as a major underlying factor for the carcinogenicity of alcoholic beverages and especially for oesophageal and oral cancer. Acetaldehyde associated with alcohol consumption is regarded as 'carcinogenic to humans' (IARC Group 1), with sufficient evidence available for the oesophagus, head and neck as sites of carcinogenicity. At present, research into the mechanistic aspects of acetaldehyde-related oral cancer has been focused on salivary acetaldehyde that is formed either from ethanol metabolism in the epithelia or from microbial oxidation of ethanol by the oral microflora. This study was conducted to evaluate the role of the acetaldehyde that is found as a component of alcoholic beverages as an additional factor in the aetiology of oral cancer.
Salivary acetaldehyde levels were determined in the context of sensory analysis of different alcoholic beverages (beer, cider, wine, sherry, vodka, calvados, grape marc spirit, tequila, cherry spirit), without swallowing, to exclude systemic ethanol metabolism.
The rinsing of the mouth for 30 seconds with an alcoholic beverage is able to increase salivary acetaldehyde above levels previously judged to be carcinogenic in vitro, with levels up to 1000 μM in cases of beverages with extreme acetaldehyde content. In general, the highest salivary acetaldehyde concentration was found in all cases in the saliva 30 sec after using the beverages (average 353 μM). The average concentration then decreased at the 2-min (156 μM), 5-min (76 μM) and 10-min (40 μM) sampling points. The salivary acetaldehyde concentration depends primarily on the direct ingestion of acetaldehyde contained in the beverages at the 30-sec sampling, while the influence of the metabolic formation from ethanol becomes the major factor at the 2-min sampling point.
This study offers a plausible mechanism to explain the increased risk for oral cancer associated with high acetaldehyde concentrations in certain beverages.
A gas chromatography with flame ionization detection method (GC-FID) with direct injection, using a capillary column, was validated to determine ethanol, acetaldehyde, methanol, and acetone in different human matrices, such as whole blood, vitreous humour, and urine, with clinical and forensic interest. This method was also employed to quantify these compounds in cell culture medium, thus being useful in basic research. A good peak resolution was achieved, with linear correlation between concentration and peak areas for all the compounds in all the matrices. The inter- and intra-day precisions of the method were always under 15% and 10%, respectively. The accuracy of the method, calculated as the percentage of the target concentration, was within the acceptable limits. The obtained limits of detection were below 0.85 mg/L for acetaldehyde and below 0.75 mg/L for the other considered compounds. The small injection volume and the high split ratios applied, allied to the high performance of the GC column, resulted in very good peak resolution and high sensitivities. This method is easy to perform, making it suitable for the routine of clinical biochemistry and forensic laboratories.
Chronic ethanol ingestion leads to an enhanced risk of upper gastrointestinal tract cancer. Although many hypotheses for the tumor promoting effect of alcohol exist, the pathogenetic mechanisms remain unclear since alcohol in itself is not carcinogenic. Acetaldehyde, the first metabolite of ethanol, has been shown to have multiple mutagenic effects and to be carcinogenic to animals. Previous research has revealed that acetaldehyde can be formed from ethanol via microbial alcohol dehydrogenase. Thus, at least part of the proposed tumorigenic effect of ethanol may be linked to local production of acetaldehyde from ethanol by oral microflora. In this study we demonstrate the production of marked amounts of acetaldehyde in saliva after ingestion of moderate amounts of ethanol. Considerable inter individual variation in acetaldehyde production capacity is also shown. In vivo acetaldehyde production is significantly reduced after a 3-day use of an antiseptic mouthwash (chlorhexidine). In vitro acetaldehyde production was shown to be linear in time, inhibited by 4-methylpyrazole and it could not be saturated under ethanol conditions that are relevant in vivo. There was a significant positive correlation between salivary acetaldehyde production in vitro and in vivo. We conclude, that the microbial formation of acetaldehyde in saliva could be one explanation for the tumor promoting effect of ethanol on the upper gastrointestinal tract. Moreover, this may support the epidemiological finding, that poor oral hygiene is an independent risk factor for oral cavity cancer.
A sensitive, fast, simple, and high-throughput enzymatic method for the quantification of ethanol in whole blood (blood) on Hitachi 917 is presented. Alcohol dehydrogenase (ADH) oxidizes ethanol to acetaldehyde using the coenzyme nicotinamide adenine dinucleotide (NAD), which is concurrently reduced to form NADH. Method development was performed with the aid of factorial design, varying pH, and concentrations of NAD+ and ADH. The linear range increased and reaction end point decreased with increasing NAD+ concentration and pH. The method was linear in the concentration range 0.0024-0.4220 g/dL. The limits of detection and quantification were 0.0007 g/dL and 0.0024 g/dL, respectively. Relative standard deviations for the repeatability and within-laboratory reproducibility were in the ranges 0.7-5.7% and 1.6-8.9%, respectively. The correlation coefficient when compared with headspace gas chromatography-flame ionization detection methods was 0.9903. Analysis of authentic positive blood specimens gave results that were slightly lower than those of the reference method.
In a previous work, an automated alcohol dehydrogenase method for the quantification of ethanol in whole blood (blood) specimens
was presented. In the present work the application of the method to urine specimens has been investigated. Also, method robustness
to routine analysis of urine and blood specimens during a period of eight months is shown. The limits of detection and quantification
for urine were 0.0012 g/dL and 0.0042 g/dL, respectively. Relative standard deviations for the repeatability and within-laboratory
reproducibility were in the ranges 1.4–4.1% and 1.8–4.6%, respectively. The method was compared with two headspace gas chromatography-flame
ionization detection methods using authentic forensic urine specimens (n = 305) and blood specimens (n = 3186). Passing-Bablok regression for the concentration range 0.01–0.48 g/dL (urine) and 0.002–0.40 g/dL (blood) showed
a statistically significant difference, for urine y = 0.9313 (0.9250 – 0.9377)x + 0.0038 (0.0029–0.0044) and for blood y = 0.9493 (0.9491 – 0.9495)x + 0.0032 (0.00318–0.00323), at 95% confidence level. The results of the external quality control specimens were in accordance
with the reported theoretical concentrations.
Analytical method validation is the systematic process of establishing that an analytical method is acceptable for its intended purpose. In general the developer or user of the method generates evidence on specificity, linearity range, accuracy, precision, detection limit, quantitation limit, ruggedness and robustness of the method for regulatory submissions or in-house application. The iterative process of method development and validation has a direct impact on the quality of the above data. Such validated analytical methods for qualitative or quantitative testing of drug molecules assume greater importance when they are employed to generate quality and safety compliance data during development and post-approval of drug products. The present paper aims to discuss salient points of the analytical method development and validation cycle. It also attempts to compare and summarize guidelines issued by different agencies for validation of analytical methods used for analysis of drug substances in the pure form and in pharmaceutical formulations.
A fluorometric-coupled reaction for the accurate and rapid determination malate, citrate, pyruvate and oxaloacetate is presented. The method was found useful for an accurate and rapid determination of these metabolites in low volumes of milk, yogurt, apple and lemon juice and wines without considerable pretreatment. In particular, this method was found valuable in characterising the outcome of maloactic acid fermentation (MLF) in wine and outlined for the first time fundamental differences in MLF between red and white wines. Thus, this method has merit in analysing these substances in heterogeneous, opaque and colorful foods.
The consumption of ethanol is known to increase the likelihood of oral cancer. In addition, there has been a growing concern about possible association between long term use of ethanol-containing mouthwashes and oral cancer. Acetaldehyde, known to be a carcinogen, is the first metabolite of ethanol and it can be produced in the oral cavity after consumption or exposure to ethanol. This paper reports on the development of a gas-diffusion flow injection method for the online determination of salivary acetaldehyde by its colour reaction with 3-methyl-2-benzothiazolinone hydrazone (MBTH) and ferric chloride. Acetaldehyde samples and standards (80μL) were injected into the donor stream containing NaCl from which acetaldehyde diffused through the hydrophobic Teflon membrane of the gas-diffusion cell into the acceptor stream containing the two reagents mentioned above. The resultant intense green coloured dye was monitored spectrophotometrically at 600nm. Under the optimum working conditions the method is characterized by a sampling rate of 9h(-1), a linear calibration range of 0.5-15mgL(-1) (absorbance=5.40×10(-2) [acetaldehyde, mgL(-1)], R(2)=0.998), a relative standard deviation (RSD) of 1.90% (n=10, acetaldehyde concentration of 2.5mgL(-1)), and a limit of detection (LOD) of 12.3μgL(-1). The LOD and sampling rate of the proposed method are superior to those of the conventional gas chromatographic (GC) method (LOD=93.0μgL(-1) and sampling rate=4h(-1)). The reliability of the proposed method was illustrated by the fact that spiked with acetaldehyde saliva samples yielded excellent recoveries (96.6-101.9%), comparable to those obtained by GC (96.4-102.3%) and there was no statistically significant difference at the 95% confidence level between the two methods when non-spiked saliva samples were analysed.
Replacement of NAD by thio-NAD and measurement at 405 rather than 340 nm can be used in the determination of lactose and galactose. These modifications allow microplate-readers rather than UV spectrophotometers to be used in the assays. r
A fluorometric-coupled reaction for the accurate and rapid determination d- and l-lactate and lactose, galactose in foods is presented. The method was found useful for an accurate determination of these metabolites in heterogeneous, opaque and colourful foods without pretreatments. Example for the determination of lactose, galactose, d- and l-lactate in milk, and yogurts and d- and l-lactate in milk, wine and beer is provided. Unexpectedly, we found that the composition of some commercial bio-yogurts produced in Israel is not consistent with the classical definition of yogurts. Thus, this method offers rapid and accurate methodology, which should be particularly valuable in food quality control.
The aim of this study was to evaluate the distribution of xanthine oxidoreductase (XOR) and its two forms, xanthine oxidase (XO) and xanthine dehydrogenase (XD), in milk fractions. XOR associated with milk phospholipid membranes was found to be distributed among an intra-membranous pool in which it takes the form of a mixture of XO and XD, with a clear predominance of XD, and a free pool of XO, of which 33% is found in the outer surface of milk fat globule membrane, 20.5% in the outer surface of whey membrane particles, and the remaining 46.7% in apparent solution. The inner-membrane XOR may play a nonenzymatic role in fat secretion, whereas extra-membranous XO is freely available for a role in the innate gland immune system and may affect milk quality.
The aim of this study was to explore oral exposure to carcinogenic (group 1) acetaldehyde after single sips of strong alcoholic beverages containing no or high concentrations of acetaldehyde. Eight volunteers tasted 5 ml of ethanol diluted to 40 vol.% with no acetaldehyde and 40 vol.% calvados containing 2400 μM acetaldehyde. Salivary acetaldehyde and ethanol concentrations were measured by gas chromatography. The protocol was repeated after ingestion of ethanol (0.5 g/kg body weight). Salivary acetaldehyde concentration was significantly higher after sipping calvados than after sipping ethanol at 30s both with (215 vs. 128 μmol/l, p<0.05) and without (258 vs. 89 μmol/l, p<0.05) alcohol ingestion. From 2 min onwards there were no significant differences in the decreasing salivary acetaldehyde concentration, which remained above the level of carcinogenicity still at 10 min. The systemic alcohol distribution from blood to saliva had no additional effect on salivary acetaldehyde after sipping of the alcoholic beverages. Carcinogenic concentrations of acetaldehyde are produced from ethanol in the oral cavity instantly after a small sip of strong alcoholic beverage, and the exposure continues for at least 10 min. Acetaldehyde present in the beverage has a short-term effect on total acetaldehyde exposure.
We compared results obtained with four commercially available kits for the enzymatic (alcohol dehydrogenase) determination of ethanol in blood, in a practical test in which standardized blood samples were used. Each of the kits yielded reliable results with acceptable reproducibility. A tendency to record slightly lower values in the intermediate and high alcohol range is most likely related to incomplete (inhomogeneous) deproteinization. Blood samples containing ethanol plus various concentrations of methanol and isopropanol were analyzed to evaluate the specificity of assays. Highly toxic blood concentrations of methanol (1.5 g/liter) increased apparent ethanol values only insignificantly, but even small concentrations of isopropanol (0.5 g/liter) interfered in all kits to different but substantial extents. The specific technical characteristics of the kits, their advantages and disadvantages are discussed. Costs are compared for analysis of small numbers of samples.
In this review selected GC methods used in the analysis of alcohol are discussed. The separation of alcohol from biologic
samples is achieved by means of head space, distillation or extraction. Methods which eliminate sample preparation by direct
injection of blood or urine are also described. Newly developed breath methods utilizing gas chromatography are cited. Recent
advances in automation in the analysis of alcohol by GC are also elaborated.
Selected chemical, biochemical and instrumental methods, excluding gas chromatographic methods, for the analysis of ethanol in blood, breath and urine samples are briefly described. This review is limited solely to methodologies; no attempt being made to discuss interpretation of alcohol levels. Special emphasis is placed on the review of breath methods because of their widespread use by law enforcement agencies.
Little is known about the possible existence of endogenous acetaldehyde in human blood. This has partly been due to analytical difficulties preventing accurate determination of blood acetaldehyde levels with and without the presence of ethanol. In the present study the possible existence of endogenous acetaldehyde in human blood was investigated with headspace gas chromatography using three different procedures for the treatment of samples: (1) no treatment of whole blood, cells, or plasma (direct headspace method), (2) hemolysation, and (3) perchloric acid (PCA) precipitation, prior to the headspace determinations. In in vitro experiments, with the direct headspace and the hemolysation methods, higher acetaldehyde peaks were obtained depending on the headspace incubation time, temperature and ethanol concentration. Both methods displayed about the same values of acetaldehyde in blood cells, ranging between 0 and 40 microM, at incubation times between 15 and 180 min, an incubation temperature of 65 degrees C, and ethanol concentrations less than 5 microM. Less acetaldehyde formation (0-15 microM) was obtained with PCA precipitates of whole blood and cell components. Very low acetaldehyde levels (0-1 microM) were obtained in the supernatants without precipitates from either whole blood or cells after headspace equilibration. Substantially less acetaldehyde was formed in plasma preparations than with whole blood and cell fractions. In human experiments, the disturbance of endogenous or exogenous ethanol was minimized by separating and washing the blood cells followed by PCA treatment. No differences in acetaldehyde concentrations were observed in blood samples taken before, during, or after ethanol intoxication (1.5 g/kg dose) of four healthy non-alcoholic volunteers.(ABSTRACT TRUNCATED AT 250 WORDS)
Alcoholic beverage consumption is classified as a known human carcinogen, causally related to an increased risk of cancer of the upper gastrointestinal tract. The formation of acetaldehyde from ethanol metabolism seems to be the major mechanism underlying this effect. Acetaldehyde is carcinogenic in rodents and causes sister chromatid exchanges and chromosomal aberrations in human cells. The best-studied DNA adduct from acetaldehyde is N(2)-ethyl-2'-deoxyguanosine, which is increased in liver DNA obtained from ethanol-treated rodents and in white blood cells obtained from human alcohol abusers. However, the carcinogenic relevance of this adduct is unclear in view of the lack of evidence that it is mutagenic in mammalian cells. A different DNA adduct, 1,N(2)-propano-2'-deoxyguanosine (PdG), can also be formed from acetaldehyde in the presence of histones and other basic molecules. PdG has been shown to be responsible for the genotoxic and mutagenic effects of crotonaldehyde. The PdG adduct can exist in either of two forms: a ring-closed form or a ring-opened aldehyde form. Whereas the ring-closed form is mutagenic, the aldehyde form can participate in the formation of secondary lesions, including DNA-protein cross-links and DNA interstrand cross-links. The formation of these types of complex secondary DNA lesions resulting from PdG may explain many of the observed genotoxic effects of acetaldehyde described above. Repair of PdG and its associated adducts is complex, involving multiple pathways. Inherited variation in the genes encoding the proteins involved in the repair of PdG and its secondary adducts may contribute to susceptibility to alcoholic beverage-related carcinogenesis.
Approximately 3.6% of cancers worldwide derive from chronic alcohol drinking, including those of the upper aerodigestive tract, the liver, the colorectum and the breast. Although the mechanisms for alcohol-associated carcinogenesis are not completely understood, most recent research has focused on acetaldehyde, the first and most toxic ethanol metabolite, as a cancer-causing agent. Ethanol may also stimulate carcinogenesis by inhibiting DNA methylation and by interacting with retinoid metabolism. Alcohol-related carcinogenesis may interact with other factors such as smoking, diet and comorbidities, and depends on genetic susceptibility.
It has long been suggested that some of the neuropharmacological, neurochemical and behavioural effects of ethanol are mediated by its first metabolite, acetaldehyde. In spite of the well documented psychoactivity of acetaldehyde, the precise role of this compound in alcohol abuse remains a matter of intense debate among scientists devoted to the study of alcoholism. Very frequently, the main drawback has been related to the presence of adequate levels of acetaldehyde or its derivatives inside the brain after ethanol ingestion. Since penetration into the central nervous system from blood of peripherically derived acetaldehyde is very low due to the high aldehyde dehydrogenase activity at the blood-brain barrier, several authors called into question the acetaldehyde implication in the toxicity and neurobehavioral effects of ethanol. The confirmation in several laboratories of the existence of enzymatic mechanisms of ethanol oxidation in the brain has revitalized the old theories supporting the acetaldehyde contribution to alcohol abuse and alcoholism. In this paper, we review current data on the brain metabolism of ethanol. We focused on the description of the enzymatic mechanisms involved in this metabolic process, reviewing the constitutive expression, catalytic activity and inhibition and inducibility of the enzymes involved in brain ethanol metabolism. We also analyze old and recent data on their regional distribution and cellular localization in the central nervous system, with special reference to the mesocorticolimbic system, a dopaminergic brain pathway that plays an important role in drug and ethanol reinforcement.
Acetaldehyde is a volatile compound naturally found in alcoholic beverages, and it is regarded as possibly being carcinogenic to humans (IARC Group 2B). Acetaldehyde formed during ethanol metabolism is generally considered as a source of carcinogenicity in alcoholic beverages. However, no systematic data is available about its occurrence in alcoholic beverages and the carcinogenic potential of human exposure to this directly ingested form of acetaldehyde outside ethanol metabolism. In this study, we have analysed and evaluated a large sample collective of different alcoholic beverages (n=1,555). Beer (9+/-7 mg/l, range 0-63 mg/l) had significantly lower acetaldehyde contents than wine (34+/-34 mg/l, range 0-211 mg/l), or spirits (66+/-101 mg/l, range 0-1,159 mg/l). The highest acetaldehyde concentrations were generally found in fortified wines (118+/-120 mg/l, range 12-800 mg/l). Assuming an equal distribution between the beverage and saliva, the residual acetaldehyde concentrations in the saliva after swallowing could be on average 195 microM for beer, 734 microM for wine, 1,387 microM for spirits, or 2,417 microM for fortified wine, which are above levels previously regarded as potentially carcinogenic. Further research is needed to confirm the carcinogenic potential of directly ingested acetaldehyde. Until then, some possible preliminary interventions include the reduction of acetaldehyde in the beverages by improvement in production technology or the use of acetaldehyde binding additives. A re-evaluation of the 'generally recognized as safe' status of acetaldehyde is also required, which does not appear to be in agreement with its toxicity and carcinogenicity.
Dietary sugars: Chemistry, analysis, function and effects. Food and nutritional components in focus pages
- N Silanikove
- F Shapiro
Silanikove, N., & Shapiro, F. (2012). Combined assays for lactose and galactose by
enzymatic reactions. In V. R. Preedy (Ed.), Dietary sugars: Chemistry, analysis,
function and effects. Food and nutritional components in focus pages
(pp. 395-404). Cambridge, UK: Royal Soc. Chem.