Validation of a modified alcohol dehydrogenase assay for ethanol determination

Ishmayana, Safri; Fadhlillah, Muhammad; Kristia, Yogi Y.; Budiman, Harry

doi:10.5267/j.ccl.2015.1.001

Cited by 8 publications

(7 citation statements)

References 7 publications

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“…1,10 Therefore, accurate assessment of potential tolerance mutants requires correcting for initial resistance phenotypes. As mentioned above, a clear indication of a primary tolerance mutant is that compared to the control, the mutant has a tolerance phenotype but no initial resistance phenotype (as is the case for mutants PQBP1 2-10 and sgg [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21] , Figure 9A). However, when a mutant has both an initial resistance and tolerance phenotype, it is less clear whether it is a primary tolerance mutant.…”

Section: Discussionmentioning

confidence: 82%

“…Residuals (y-axis) in the XY plot (A) indicate how far from the inverse correlation curve the data points lie, while the frequency plot (B) shows a normal distribution of the residuals centered around 0 (mean = 0.004). Some potential strong, primary tolerance mutants are highlighted (Arf6, Efa6, CASK, NMDAR, Gprk2, No66, PQBP1 2-10 and sgg[5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21] ) as well as GNMT (yellow), which is not a primary tolerance mutant. (C) Example of how to calculate residuals using the equation y(exp) = À0.56 x À 0.23 with GNMT, Arf6 and Gprk2 data, where y(exp) stands for the tolerance effect size (y) expected.…”

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confidence: 99%

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Large analysis of genetic manipulations reveals an inverse correlation between initial alcohol resistance and rapid tolerance phenotypes

Chvilicek,

Seguin,

Lathen

et al. 2024

Genes Brain and Behavior

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Tolerance occurs when, following an initial experience with a substance, more of the substance is required subsequently to induce identical behavioral effects. Tolerance is not well‐understood, and numerous researchers have turned to model organisms, particularly Drosophila melanogaster, to unravel its mechanisms. Flies have high translational relevance for human alcohol responses, and there is substantial overlap in disease‐causing genes between flies and humans, including those associated with Alcohol Use Disorder. Numerous Drosophila tolerance mutants have been described; however, approaches used to identify and characterize these mutants have varied across time and labs and have mostly disregarded any impact of initial resistance/sensitivity to ethanol on subsequent tolerance development. Here, we analyzed our own, as well as data published by other labs to uncover an inverse correlation between initial ethanol resistance and tolerance phenotypes. This inverse correlation suggests that initial resistance phenotypes can explain many ‘perceived’ tolerance phenotypes, thus classifying such mutants as ‘secondary’ tolerance mutants. Additionally, we show that tolerance should be measured as a relative increase in time to sedation between an initial and second exposure rather than an absolute change in time to sedation. Finally, based on our analysis, we provide a method for using a linear regression equation to assess the residuals of potential tolerance mutants. These residuals provide predictive insight into the likelihood of a mutant being a ‘primary’ tolerance mutant, where a tolerance phenotype is not solely a consequence of initial resistance, and we offer a framework for understanding the relationship between initial resistance and tolerance.

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Section: Discussionmentioning

confidence: 82%

mentioning

confidence: 99%

Large analysis of genetic manipulations reveals an inverse correlation between initial alcohol resistance and rapid tolerance phenotypes

Chvilicek,

Seguin,

Lathen

et al. 2024

Genes Brain and Behavior

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“…Then 25 μL of supernatant was mixed with 300 μL semicarbazide buffer (3.3% tetrasodium pyrophosphate, 0.84% semicarbazide hydrochloride and 0.16% Glycine), 25 μL 1.6% NAD+ and 25 μL of alcohol dehydrogenase (4000 units / mL). The solution was incubated at 40°C for 40 minutes and absorbance at 340 nm for each sample determined on a NanoDrop spectrophotometer (Ishmayana et al, 2015). We repeated the procedure at least three times for each genotype (n ≥ 3).…”

Section: Methodsmentioning

confidence: 99%

Alcohol‐Induced Behaviors Require a Subset of Drosophila JmjC‐Domain Histone Demethylases in the Nervous System

Pinzón

Reed

Shalaby

et al. 2017

Alcoholism Clin & Exp Res

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Background Long-lasting transcriptional changes underlie a number of adaptations that contribute to alcohol use disorders (AUD). Chromatin remodeling, including histone methylation, can confer distinct, long-lasting transcriptional changes, and histone methylases are known to play a role in the development of addiction. Conversely, little is known about the relevance of Jumonji (JmjC) domain-containing demethylases in AUDs. We systematically surveyed the alcohol-induced phenotypes of null mutations in all 13 Drosophila JmjC genes. Methods We used a collection of JmjC mutants, the majority of which we generated by homologous recombination, and assayed them in the Booze-o-mat to determine their naïve sensitivity to sedation and their tolerance (change in sensitivity upon repeat exposure). Mutants with reproducible phenotypes had their phenotypes rescued with tagged genomic transgenes, and/or phenocopied by nervous system-specific knock down using RNA interference (RNAi). Results Four of the 13 JmjC genes (KDM3, lid, NO66 and HSPBAP1) showed reproducible ethanol-sensitivity phenotypes. Some of the phenotypes were observed across doses, e.g. the enhanced ethanol-sensitivity of KDM3KO and NO66KO, but others were dose-dependent, such as the reduced ethanol sensitivity of HSPBAP1KO, or the enhanced ethanol tolerance of NO66KO. These phenotypes were rescued by their respective genomic transgenes in KDM3KO and NO66KO mutants. While we were unable to rescue lidk mutants, knock down of lid in the nervous system recapitulated the lidk phenotype, as was observed for KDM3KO and NO66KO RNAi-mediated knock down. Conclusion Our study reveals that the Drosophila JmjC-domain histone demethylases Lid, KDM3, NO66, and HSPBAP1 are required for normal ethanol-induced sedation and tolerance. Three of three tested of those four JmjC genes are required in the nervous system for normal alcohol-induced behavioral responses, suggesting that this gene family is an intriguing avenue for future research.

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“…Kadar etanol ditentukan dengan metode enzimatik menggunakan enzim alkohol dehidrogenase (Ishmayana et al 2015b). Sebanyak 25 µL, sampel diambil dan ditambahkan dengan 1,25 mL buffer semikarbazida dan diaduk.…”

Section: Penentuan Kadar Etanolunclassified

Peranan Ion Logam Kobalt Terhadap Kinerja Fermentasi dan Toleransi Cekaman Lingkungan Sel Ragi Saccharomyces cerevisiae

et al. 2019

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Semakin menipisnya ketersediaan bahan bakar fosil mendorong dikembangkannya energi alternatif, salah satunya adalah bioetanol. Selama proses fermentasi bioetanol, ragi Saccharomyces cerevisiae dapat terpapar berbagai cekaman lingkungan yang dapat memberikan dampak negatif bagi sel ragi. Salah satu upaya untuk meningkatkan toleransi sel ragi adalah dengan menambahkan ion logam kedalam media fermentasi. Penelitian ini bertujuan untuk mengetahui pengaruh suplementasi ion kobalt terhadap kinerja fermentasi, toleransi sel ragi terhadap cekaman lingkungan, dan terhadap morfologi sel ragi. Metode penelitian meliputi proses fermentasi yang dilaksanakan selama 120 jam dalam media yeast nitrogen base dengan 10% b/v glukosa sebagai sumber karbon dan variasi penambahan konsentrasi ion kobalt. Sampling dilakukan tiap 6 jam sekali pada 24 jam pertama dan tiap 12 jam sekali sampai jam ke-120. Kurva pertumbuhan ditentukan dengan mengukur OD600 nm sedangkan viabilitas sel ditentukan dengan pewarnaan metilen violet. Kadar glukosa dalam media fermentasi diukur dengan metode kalium ferisianida basa sedangkan kadar etanol ditentukan dengan metode enzimatik menggunakan alkohol dehidrogenase. Selain itu dilakukan penentuan ukuran sel serta tingkat toleransi sel terhadap cekaman lingkungan. Etanol paling tinggi diperoleh sebesar 2,58% pada media yang disuplementasi 40 ppm ion kobalt, lebih tinggi dibandingkan dengan kontrol yang menghasilkan bioetanol sebesar 2,20%. Hasil penelitian ini menunjukkan bahwa ion kobalt tidak memberikan pengaruh yang signifikan terhadap kinerja fermentasi sel ragi, toleransi sel ragi dalam menghadapi cekaman lingkungan dan terhadap diameter sel.

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Validation of a modified alcohol dehydrogenase assay for ethanol determination

Cited by 8 publications

References 7 publications

Large analysis of genetic manipulations reveals an inverse correlation between initial alcohol resistance and rapid tolerance phenotypes

Large analysis of genetic manipulations reveals an inverse correlation between initial alcohol resistance and rapid tolerance phenotypes

Alcohol‐Induced Behaviors Require a Subset of Drosophila JmjC‐Domain Histone Demethylases in the Nervous System

Peranan Ion Logam Kobalt Terhadap Kinerja Fermentasi dan Toleransi Cekaman Lingkungan Sel Ragi Saccharomyces cerevisiae

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