Analytical and mechanistic aspects of the electrochemical oxidation of keto steroids derivatized with phenylhydrazine, (4-nitrophenyl)hydrazine, and (2,4-dinitrophenyl)hydrazine

Bond, Alan M.; Hollenkamp, Anthony F.; Thompson, Stephen B.; Bourne, A.R.; Huf, P.A.; Watson, T.G.

doi:10.1021/ac00161a015

Cited by 15 publications

(8 citation statements)

References 23 publications

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“…We further tested activity of the enzymes on phenylhydrazine, a potent reducing agent with E° = 410 mV [47] that readily reacts with Cu(II) (Figure 2F). It has previously been described as both an inhibitor and substrate for a tyrosinase [48], ultimately resulting in an o -quinone.…”

Section: Resultsmentioning

confidence: 99%

Characterization of an Alkali- and Halide-Resistant Laccase Expressed in E. coli: CotA from Bacillus clausii

2014

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The limitations of fungal laccases at higher pH and salt concentrations have intensified the search for new extremophilic bacterial laccases. We report the cloning, expression, and characterization of the bacterial cotA from Bacillus clausii, a supposed alkalophilic ortholog of cotA from B. subtilis. Both laccases were expressed in E. coli strain BL21(DE3) and characterized fully in parallel for strict benchmarking. We report activity on ABTS, SGZ, DMP, caffeic acid, promazine, phenyl hydrazine, tannic acid, and bilirubin at variable pH. Whereas ABTS, promazine, and phenyl hydrazine activities vs. pH were similar, the activity of B. clausii cotA was shifted upwards by ∼0.5–2 pH units for the simple phenolic substrates DMP, SGZ, and caffeic acid. This shift is not due to substrate affinity (KM) but to pH dependence of catalytic turnover: The kcat of B. clausii cotA was 1 s−1 at pH 6 and 5 s−1 at pH 8 in contrast to 6 s−1 at pH 6 and 2 s−1 at pH 8 for of B. subtilis cotA. Overall, kcat/KM was 10-fold higher for B. subtilis cotA at pHopt. While both proteins were heat activated, activation increased with pH and was larger in cotA from B. clausii. NaCl inhibited activity at acidic pH, but not up to 500–700 mM NaCl in alkaline pH, a further advantage of the alkali regime in laccase applications. The B. clausii cotA had ∼20 minutes half-life at 80°C, less than the ∼50 minutes at 80°C for cotA from B. subtilis. While cotA from B. subtilis had optimal stability at pH∼8, the cotA from B. clausii displayed higher combined salt- and alkali-resistance. This resistance is possibly caused by two substitutions (S427Q and V110E) that could repel anions to reduce anion-copper interactions at the expense of catalytic proficiency, a trade-off of potential relevance to laccase optimization.

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Section: Resultsmentioning

confidence: 99%

Characterization of an Alkali- and Halide-Resistant Laccase Expressed in E. coli: CotA from Bacillus clausii

2014

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“…For method development, studies on derivatization of some hydrazines with MPA had been carried out in acidic solution [15]. It was found that DNPH is a suitable reagent.…”

Section: Resultsmentioning

confidence: 99%

“…Numerous recent researches for this analysis have been reported concerning the method sensitivity and selectivity in association with various instruments including GC, HPLC and others [6][7][8][9][10][11][12][13][14][15]. However, each of which had their advantages and disadvantages.…”

Section: Determination Of Mpa In Hormonal Contraceptive Drugsmentioning

confidence: 99%

“…From the molecular structure, MPA is a ketone steroid in which the ketone group can undergo reduction. Since the reduction is normally easily interfered by oxygen molecule dissolved in mobile phase, this may be a problem [15]. Hydrazines including 2,4-dinitrophenyl hydrazine (DNPH) have been introduced as an electroactive labeling reagent for carbonyl compounds [16].…”

Section: Determination Of Mpa In Hormonal Contraceptive Drugsmentioning

confidence: 99%

See 1 more Smart Citation

Application of HPLC Analysis of Medroxyprogesterone Acetate in Human Plasma

Chanthai¹,

Tessiri²

2012

Chromatography - The Most Versatile Method of Chemical Analysis

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“…First phenyl hydrazine undergoes oxidation to diazenyl benzene as shown in the step 1 of Scheme 2 and attach to the electrode surface as shown in step 1 of Scheme 3 followed by two electron transfer to the electrode and the oxidized phenyl hydrazine then helps the T 4 to undergo oxidation probably either one of the way (2a or 2b) as shown in Scheme 3. But when the system contains methanol (since the dilution media for sodium levothyroxine is methanol) then there is a chance of formation of methoxy benzene, hydrazine with dinitrogen as the leaving group in small amounts giving the overall reaction as shown in Scheme 2 [28][29][30][31].…”

Section: Electrocatalytic Oxidation Of Sodium Levothyroxine At Carbonmentioning

confidence: 99%

Electrocatalytic oxidation of sodium levothyroxine with phenyl hydrazine as a mediator at carbon paste electrode: A cyclic voltammetric study

Chitravathi¹,

Swamy

Chandra

et al. 2010

Journal of Electroanalytical Chemistry

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Analytical and mechanistic aspects of the electrochemical oxidation of keto steroids derivatized with phenylhydrazine, (4-nitrophenyl)hydrazine, and (2,4-dinitrophenyl)hydrazine

Cited by 15 publications

References 23 publications

Characterization of an Alkali- and Halide-Resistant Laccase Expressed in E. coli: CotA from Bacillus clausii

Characterization of an Alkali- and Halide-Resistant Laccase Expressed in E. coli: CotA from Bacillus clausii

Application of HPLC Analysis of Medroxyprogesterone Acetate in Human Plasma

Electrocatalytic oxidation of sodium levothyroxine with phenyl hydrazine as a mediator at carbon paste electrode: A cyclic voltammetric study

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