Monitoring steady production of 1,3‐propanediol during bioprospecting of glycerol‐assimilating soil microbiome using dye‐based pH‐stat method

Kinetics and mechanism of hydrolysis of acetylthiocholine by the enzyme butyrylcholine esterase was studied. The spectrophotometric Ellman’s method and potentiometric pH-stat method were used for continuous determination of the actual concentration of the products thiocholine and acetic acid in the reaction mixture. The validity of the Michaelis-Menten (Briggs-Haldane) equation in the whole course of the reaction under used conditions was proved. The corresponding kinetics parameters (Vm and KM) were calculated from the obtained dependences of concentration of thiocholine or acetic acid vs. time and compared. From this comparison the deciding kinetic role of the step producing thiocholine was derived. The values of initial molar concentration of the enzyme and of the rate constants of the kinetic model were estimated.

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Bicarbonate Transport Systems in the Intestine of the Seawater EEL

Journal of Experimental Biology ◽

10.1242/jeb.150.1.381 ◽

1990 ◽

Vol 150 (1) ◽

pp. 381-394 ◽

Cited By ~ 8

Author(s):

MASAAKI ANDO ◽

M. V. SUBRAMANYAM

Keyword(s):

Latent Period ◽

Water Transport ◽

Basolateral Membrane ◽

Transport Systems ◽

Bicarbonate Transport ◽

Ph Stat ◽

Stat Method

Utilizing a pH-stat method, the rates of mucosal and serosal alkalinization were measured separately in the seawater eel intestine. These two rates were dependent on contralateral HCO3− concentration and were inhibited by contralateral application of DIDS, an inhibitor of HCO3− transport, indicating that the mucosal and serosal alkalinization are due to HCO3− secretion and absorption, respectively. The mucosal alkalinization was enhanced after inhibiting Na+/K+/Cl− cotransport by treatment with bumetanide, furosemide or Ba2+, with a latent period of more than lOmin, suggesting that HCO3− absorption from mucosa to serosa depends on Na+/K+/Cl− cotransport. The serosal alkalinization caused by HCO3− absorption was completely abolished after mucosal application of bumetanide. After pretreatment with bumetanide, mucosal omission of Cl− halved the enhanced rate of mucosal alkalinization, and Na+ omission had no effect on it; this indicates that the exit of HCO3− into the lumen depends on luminal Cl−, i.e. on the existence of the usual C1−/HCO3− exchange on the brushborder membrane. When serosal Na+ was removed under the same conditions, mucosal alkalinization was reduced, indicating that HCO3− entry from the serosal fluid depends on Na+. Serosal omission of Cl− did not reduce mucosal alkalinization. In addition, serosal alkalinization was enhanced by serosal removal of Na+ but not of Cl−. These results suggest that there is a Na+/HCO3− cotransport on the basolateral membrane. A possible model for HCO3− transport systems in the seawater eel intestine is proposed, and a possible role for these transport systems is discussed in relation to Na+, Cl− and water transport.

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Lipase activity measured in serum by a continuous-monitoring pH-Stat technique--an update.

Clinical Chemistry ◽

10.1093/clinchem/35.8.1688 ◽

1989 ◽

Vol 35 (8) ◽

pp. 1688-1693 ◽

Cited By ~ 21

Author(s):

N W Tietz ◽

J R Astles ◽

D F Shuey

Keyword(s):

Olive Oil ◽

Continuous Monitoring ◽

Hydroxypropyl Methylcellulose ◽

Reference Interval ◽

Sample Volume ◽

Standard Curve ◽

Activity Measurements ◽

Ph Stat ◽

Stat Method ◽

Assay Conditions

Abstract Using recent knowledge regarding the roles of colipase, bile acids, Ca2+, and emulsifiers, we optimized a previously published pH-Stat method for lipase (EC 3.1.1.3) activity measurements. The recommended assay conditions are: olive oil/triolein, 100 mL/L; sodium glycocholate, 35 mmol/L; Ca2+, 8.5 mmol/L; and colipase, 6.0 mg/L. The sample volume is 0.10 mL, the reaction pH 9.0, the temperature 30 degrees C, and the concentration of titrant 15 mmol/L. Hydroxypropyl methylcellulose, 20 g/L, replaces acacia as emulsifier to avoid inhibition by excess Ca2+. The standard curve is linear to greater than 4566 U/L. The reference interval with olive oil as substrate is 30-235 U/L. Lipase activities with triolein substrate are 9.9% greater than with olive oil. Interference by pancreatic carboxylesterase (EC 3.1.1.1) activity is inhibited by incubating the sample with diisopropylfluorophosphate. Results correlate well with those by the optimized SingleVial method of Boehringer Mannheim Diagnostics (r = 0.997) and the immunochemical assay of Beckman Instruments, Inc. (r = 0.995). Correlation with the aca method (E.I. DuPont de Nemours & Company) is less satisfactory (r = 0.892), probably owing to lack of colipase in the latter method.

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Use of Silica Based Materials as Modulators of the Lipase Catalyzed Hydrolysis of Fats under Simulated Duodenal Conditions

Nanomaterials ◽

10.3390/nano10101927 ◽

2020 ◽

Vol 10 (10) ◽

pp. 1927

Author(s):

Sara Muñoz-Pina ◽

Pedro Amorós ◽

Jamal El Haskouri ◽

Ana Andrés ◽

José V. Ros-Lis

Keyword(s):

Partial Least Square ◽

Least Square ◽

Partial Least Square Regression ◽

Fat Globules ◽

Silica Materials ◽

Functionalized Materials ◽

Ph Stat ◽

Surface Functionalizations ◽

Fat Hydrolysis ◽

Stat Method

The effect of silica materials and their functionalization in the lipase catalyzed fat hydrolysis has been scarcely studied. Fifteen silica materials were prepared and their effect on the fat hydrolysis was measured, under simulated duodenal conditions, using the pH-stat method. The materials are composed of the combination of three supports (Stöber massive silica nanoparticles, Stöber mesoporous nanoparticles and UVM-7) and four surface functionalizations (methyl, trimethyl, propyl and octyl). In addition, the non-functionalized materials were tested. The functional groups were selected to offer a hydrophobic character to the material improving the interaction with the fat globules and the lipase. The materials are able to modulate the lipase activity and their effect depending on the support topology and the organic covering, being able to increase or reduce the fat hydrolysis. Depending of the material, relative fat hydrolysis rates of 75 to 140% in comparison with absence of the material were obtained. The results were analyzed by Partial Least Square Regression and suggest that the alkyl modified mesopores are able to improve the fat hydrolysis, by contrast the non-porous nanoparticles and the textural pores tend to induce inhibition. The effects are more pronounced for materials containing long alkyl chains and/or in absence of taurodeoxycholate.

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Physiological changes in human erythrocyte cholinesterase as measured with the "pH-stat".

Clinical Chemistry ◽

10.1093/clinchem/34.10.2133 ◽

1988 ◽

Vol 34 (10) ◽

pp. 2133-2135 ◽

Cited By ~ 5

Author(s):

J A Garcia-López ◽

M Monteoliva

Keyword(s):

Human Erythrocyte ◽

Cholinesterase Activity ◽

Human Erythrocytes ◽

Physiological Changes ◽

Individual Values ◽

Blood Samples ◽

Age Related ◽

Population Average ◽

Ph Stat ◽

Stat Method

Abstract Cholinesterase activity in human erythrocytes was determined in 1903 blood samples by the "pH-stat" method. Differences in activity were examined as a function of sex, age, and pregnancy. Reliability intervals for the population average and approval or normality intervals for individual values were established. Sex- and age-related differences were very significant.

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Factors affecting lipase digestibility of emulsified lipids using an in vitro digestion model: Proposal for a standardised pH-stat method

Food Chemistry ◽

10.1016/j.foodchem.2010.11.027 ◽

2011 ◽

Vol 126 (2) ◽

pp. 498-505 ◽

Cited By ~ 187

Author(s):

Yan Li ◽

Min Hu ◽

David Julian McClements

Keyword(s):

In Vitro Digestion ◽

Factors Affecting ◽

Ph Stat ◽

Stat Method

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Analysis of progress curves. Rate law of pyruvate kinase type I from Escherichia coli

Biochemical Journal ◽

10.1042/bj1890421 ◽

1980 ◽

Vol 189 (3) ◽

pp. 421-433 ◽

Cited By ~ 34

Author(s):

M Markus ◽

T Plesser ◽

A Boiteux ◽

B Hess ◽

M Malcovati

Keyword(s):

Escherichia Coli ◽

Pyruvate Kinase ◽

Regulatory Region ◽

Conformational State ◽

Type I ◽

Allosteric Site ◽

Rate Law ◽

Ph Stat ◽

Stat Method ◽

Concentration Space

Progress curves of the reaction catalysed by pyruvate kinase from Escherichia coli K12, designed to cover the four-dimensional concentration space of phosphoenolpyruvate, ADP, Mg2+ and ATP in the regulatory region, were recorded with the pH-stat method (pH 7.0 and 25 degrees C). Additional initial-rate measurement were performed to assess specific points. Two methods for the evaluation of progress curves were used: fitting the rate law to the rates obtained from the tangents of the progress curves and fitting the integrated rate law directly to the curves. Two models, both extensions of the concerted model given by Monod, Wyman & Changeux [(1965) J. Mol. Biol. 12, 88–118] with four protomers, could be fitted to the data within the experimental error. Model discrimination in favour of one of these models was possible by proper experimental design. In the selected model one conformational state of the enzyme forms the active complex. The active site of a second conformational state forms abortive complexes with Mg2+, causing strong inhibition at high Mg2+ concentrations. In the absence of ligands, most of the enzyme is in a third state that binds ATP at an allosteric site.

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