Development of Control Material for Exhaled Breath-Alcohol Testing and its Application.

Background: Breath analyser tests are used worldwide to obtain proof of alcohol intoxication and often used in the conviction of traffic violators. These tests are conducted to quickly and painlessly determine the existing concentration of alcohol in arterial blood by measuring the amount of ethanol in exhaled breath, which can be identified with an electrochemical sensor.At present, the calibration and maintenance of analysers used for these tests are typically performed regularly but lack quality control. Consequently, test results may not be accurate because of calibration deterioration.The aim of this study was to develop and evaluate the uncertainty of control materials used in breath-alcohol testing at the Bangkok Metropolitan Police Station. Material and methods: Ethyl alcohol (99.99%; Certified Reference Material grade) diluted at three different concentrations was kept under design conditions. The concentrations were 28, 67, and 134 mg/dL, determined by performing headspace gas chromatography, and the uncertainty was set as ±1.3925, ±2.8736, and ±1.8231 mg/dL (±4.97%, ±4.29%, and ±2.72% for the concentrations, respectively), as per ISO Guide 35:2017. Results: The total error percentages of the developed control materials were 4.97%, 4.29%, and 2.72% for concentrations of 28, 67, and 134 mg/dL, respectively. Each concentration of the materials was tested by using measurements from 70 breath-alcohol analysers belonging to the Bangkok Metropolitan Police Station. Conclusion: These control materials are applicable to quality assurance and standards tests and may help to ensure the accuracy of breath-alcohol testing in the future.

Evaluation of emerging inflammatory markers for predicting oxygen support requirement in COVID-19 patients.

Coronavirus disease 2019 (COVID-19), a highly contagious pathogenic viral infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread rapidly and remains a challenge to global public health. COVID-19 patients manifest various symptoms from mild to severe cases with poor clinical outcomes. Prognostic values of novel markers, including neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR) and C-reactive protein to lymphocyte ratio (CLR) calculated from routine laboratory parameters have recently been reported to predict severe cases; however, whether this investigation can guide oxygen therapy in COVID-19 patients remains unclear. In this study, we assessed the ability of these markers in screening and predicting types of oxygen therapy in COVID-19 patients. The retrospective data of 474 COVID-19 patients were categorized into mild and severe cases and grouped according to the types of oxygen therapy requirement, including noninvasive oxygen support, high-flow nasal cannula and invasive mechanical ventilator. Among the novel markers, the ROC curve analysis indicated a screening cutoff of CRP ≥ 30.0 mg/L, NLR ≥ 3.0 and CLR ≥ 25 in predicting the requirement of any type of oxygen support. The NLR and CLR with increasing cut-off values have discriminative power with high accuracy and specificity for more effective oxygen therapy with a high-flow nasal cannula (NLR ≥ 6.0 and CLR ≥ 60) and mechanical ventilator (NLR ≥ 8.0 and CLR ≥ 80). Our study thus identifies potential markers to differentiate the suitable management of oxygen therapy in COVID-19 patients at an earlier time for improving disease outcomes with limited respiratory support resources.

Genotoxic Effects of Lead and Their Impact on the Expression of DNA Repair Genes.

Exposure to lead (Pb) continues to be a significant worldwide problem. Pb is a highly poisonous heavy metal affecting several organ systems in the body. Although Pb has been shown to be genotoxic to experimental animals and humans, the underlying mechanisms are still not understood. An indirect mechanism related to the inhibition of DNA repair systems by Pb has been suggested. Heavy metals can interfere with the activities of several proteins and gene expressions. Recent studies gathered in this review article demonstrated an altered expression of DNA repair genes due to Pb toxicity. However, their findings are conflicting. Furthermore, the interaction of Pb and epigenetic mechanisms regulating gene expression may have a crucial role in the inhibition of DNA repair systems. Therefore, additional studies are needed to evaluate these findings and to obtain a complete picture of the genotoxic properties of Pb and the underlying mechanisms that may have a crucial role in carcinogenesis.

Induction of genotoxicity and mutagenic potential of heavy metals in Thai occupational workers.

Heavy metals are widely used in many industries in Thailand and found in the environment. Occupational exposure to heavy metals is often chronic and caused by environmental contaminations, potentially leading to mutations and cancer. Although the genotoxic effects of occupational exposure to multiple heavy metals have been extensively studied, the findings regarding their genotoxicity are conflicting. In this study, we focused on investigating the genotoxic effects of certain heavy metals mixtures, including lead (Pb), copper (Cu), zinc (Zn), and tin (Sn), to which workers are exposed in the manufacturing industry. The cytokinesis-blocked micronucleus (CBMN) assay in peripheral blood lymphocytes was performed, and DNA damage was assessed by measuring tumour-associated protein levels and 8-hydroxy-2'-deoxyguanosine (8-OHdG) generated by oxidative stress that causes cytotoxicity. The occupational exposure group included 110 workers exposed to heavy metal mixtures and 105 matched control subjects. We found statistically significant differences in the blood Pb, Sn, and Cu levels between the exposed workers and the control subjects (p < 0.001). Analysis of micronuclei (MN) in peripheral blood lymphocytes revealed a significantly increased frequency of MN in exposed workers compared with that in control subjects (p<0.05). Non-smoking exposed workers were selected for 8-OHdG formation and mutant p53 tests, and significant differences in the mean plasma 8-OHdG concentration (p < 0.001) were found between the occupational exposure and the control group, but no differences were found in the levels of mutant p53. Thus, chronic exposure to different heavy metals causes genotoxic effects in humans. Furthermore, the CBMN assay and 8-OHdG formation can be used as surrogate biomarkers to identify and monitor groups with higher carcinogenic risk in the early stages of toxicity. In summary, our results indicate that mixtures of heavy metals (Pb, Sn, and Cu) in manufacturing industries pose an elevated health risk due to DNA damage.

Correlation of 25-Hydroxyvitamin D Levels in Serum vs. Breastmilk in Vitamin D-Supplementation Breastfeeding Women during Lactation: Randomized Double Blinded Control Trial.

Background: Vitamin D deficiency in pregnancy and lactation increases the risk of adverse perinatal outcomes; however, although Vitamin D supplementation during pregnancy and lactation is recommended, suggested dose ranges vary. Objective: To determine whether vitamin D31,800 IU/d supplementation in lactating mothers improves their vitamin D status and breast-feeding milk. Material and Method: This was a randomized, placebo­controlled study of Thai pregnant women in their third trimester. A total of 76 Thai lactating mothers and their breast-fed infants were studied with maternal 25 Hydroxyvitamin D 25 (OH) D levels of 10-30 ng/ml determined using Liquid Chromatography Mass Spectrometry Tandem (LC-MS/MS). One group received vitamin D3 1,800 IU/d supplementation for 6 weeks, and members of the other group were given a placebo. 25 (OH) D level of colostum and 6-week serum from breast-fed milk were measured by High Performance Liquid Chromatography (HPLC). The data from the two groups were analyzed and compared. Results: The mean (±SD) maternal age was 27.16±5.13 years, and mean body mass index (BMI) was 22.29±5.08 kg/m(2). At 6 weeks, maternal 25 (OH) D levels had increased significantly in the vitamin D group (VD) 68.30±15.40 nmol/L compared to 55.15±13.57 nmol/L in the placebo group (p<0.001) measured using the Liquid Chromatography-Mass Spectrometry Tandem (LC-MS/MS) method. Breast-fed milk did not show any significant incremental change in 25 (OH) D levels measured by High Performance Liquid Chromatography (HPLC); however, the change in 25 (OH) D levels in breast milk in the VD group was significantly different from that of the placebo group (p = 0.005).` Conclusion: Vitamin D3 supplementation during lactation can increase 25 (OH) D levels in Thai breast-fed mothers. Further work is needed to determine the duration of vitamin D supplementation to normalize breast milk and breast-fed infants' 25 (OH) D level at over 75 nmol/L.

Formation of 1,2:3,4-diepoxybutane-specific hemoglobin adducts in 1,3-butadiene exposed workers.

1,3-Butadiene (BD) is an important industrial chemical that is classified as a human carcinogen. BD carcinogenicity has been attributed to its metabolism to several reactive epoxide metabolites and formation of the highly mutagenic 1,2:3,4-diepoxybutane (DEB) has been hypothesized to drive mutagenesis and carcinogenesis at exposures experienced in humans. We report herein the formation of DEB-specific N,N-(2,3-dihydroxy-1,4-butadiyl)valine (pyr-Val) in BD-exposed workers as a biomarker of DEB formation. pyr-Val was determined in BD monomer and polymer plant workers that had been previously analyzed for several other biomarkers of exposure and effect. pyr-Val was detected in 68 of 81 (84%) samples ranging from 0.08 to 0.86 pmol/g globin. Surprisingly, pyr-Val was observed in 19 of 23 administrative control subjects not known to be exposed to BD, suggesting exposure from environmental sources of BD. The mean ± SD amounts of pyr-Val were 0.11 ± 0.07, 0.16 ± 0.12, and 0.29 ± 0.20 pmol/g globin in the controls, monomer, and polymer workers, respectively, clearly demonstrating formation of DEB in humans. The amounts of pyr-Val found in this study suggest that humans are much less efficient in the formation of DEB than mice or rats at similar exposures. Formation of pyr-Val was more than 50-fold lower than has been associated with increased mutagenesis in rodents. The results further suggest that formation of DEB relative to other epoxides is significantly different in the highest exposed polymer workers compared with controls and BD monomer workers. Whether this is due to saturation of metabolic formation or increased GST-mediated detoxification could not be determined.

1,3-Butadiene: Biomarkers and application to risk assessment.

1,3-Butadiene (BD) is a known rodent and human carcinogen that is metabolized mainly by P450 2E1 to three epoxides, 1,2-epoxy-3-butene (EB), 1,2:3,4-diepoxybutane (DEB) and 1,2-epoxy-3,4-butanediol (EB-diol). The individual epoxides vary up to 200-fold in their mutagenic potency, with DEB being the most mutagenic metabolite. It is important to understand the internal formation of the individual epoxides to assign the relative risk for each metabolite and to understand the molecular mechanisms responsible for major species differences in carcinogenicity. We have conducted extensive exposure-biomarker studies on mice, rats and humans. Using low exposures that range from current occupational levels to human exposures from tobacco smoke has provided evidence that mice are very different from humans, with mice forming ∼200 times more DEB than humans at exposures of 0.1-1.5ppm BD. While no gender differences have been noted in mice and rats for globin adducts or N-7 guanine adducts, female rats and mice had 2-3-fold higher Hprt mutations and DNA-DNA cross-links, suggesting a gender difference in DNA repair. Numerous molecular epidemiology studies have evaluated globin adducts and Hprt mutations, SCEs and chromosomal abnormalities. None of the blinded studies have shown evidence of human genotoxicity at current occupational exposures and studies of globin adducts have shown similar or lower formation of adducts in females than males. If one calculates the EB dose-equivalents for the three species, mice clearly differ from rats and humans, being ∼44 and 174 times greater than rats and humans, respectively. These data provide a scientific basis for improved risk assessment of BD.

Exposure-response of 1,2:3,4-diepoxybutane-specific N-terminal valine adducts in mice and rats after inhalation exposure to 1,3-butadiene.

1,3-Butadiene (BD) is a known rodent and human carcinogen that is metabolized mainly by P450 2E1 to three epoxides, 1,2-epoxy-3-butene (EB), 1,2:3,4-diepoxybutane (DEB), and 1,2-epoxy-3,4-butanediol. The individual epoxides vary up to 200-fold in their mutagenic potency, with DEB being the most mutagenic metabolite. It is important to understand the internal formation of the individual epoxides to assign the relative risk for each metabolite and to understand the molecular mechanisms responsible for extensive species differences in carcinogenicity. This study presents a comprehensive exposure-response for the formation of the DEB-specific N,N-(2,3-dihydroxy-1,4-butadiyl)valine (pyr-Val) in mice and rats. Using nano-ultra high pressure liquid chromatography-tandem-mass spectrometry allowed analysis of pyr-Val in mice and rats exposed to BD as low as 0.1 and 0.5 ppm BD, respectively, and demonstrated significant differences in the amounts and exposure-response of pyr-Val formation. Mice formed 10- to 60-fold more pyr-Val compared to rats at similar exposures. The formation of pyr-Val increased with exposures, and the formation was most efficient with regard to formation per parts per million BD at low exposures. While formation at higher exposures appeared linear in mice, in rats formation saturated at exposures > or = 200 ppm for 10 days. In rats, amounts of pyr-Val were lower after 20 days than after 10 days of exposure, suggesting that the lifespan of rat erythrocytes may be shortened following exposure to BD. This research supports the hypothesis that the lower susceptibility of rats to BD-induced carcinogenesis results from greatly reduced formation of DEB following exposure to BD.

Accurate quantitation of standard peptides used for quantitative proteomics.

MS-based proteomics has become an indispensable tool in system biology generating a need for accurate and precise quantitation of peptide standards. The presented method utilizes ultra performance LC-MS/MS (UPLC-MS/MS) to accurately quantify peptide standards at concentrations of 0.1-10 microM. The ability for accurate quantitation of micro-molar concentrations has the advantages that quantitation can be performed routinely with high precision and the high sensitivity of the method minimizes the amounts required.