scholarly journals Increased Selectivity of Novozym 435 in the Asymmetric Hydrolysis of a Substrate with High Hydrophobicity Through the Use of Deep Eutectic Solvents and High Substrate Concentrations

Molecules ◽  
2019 ◽  
Vol 24 (4) ◽  
pp. 792 ◽  
Author(s):  
Yerko Fredes ◽  
Lesly Chamorro ◽  
Zaida Cabrera
Keyword(s):  
Substrate Concentration ◽  
Phosphate Buffer ◽  
Choline Chloride ◽  
Reaction Medium ◽  
Deep Eutectic Solvents ◽  
Novozym 435 ◽  
Chiral Drugs ◽  
High Substrate ◽  
Hydrolysis Of ◽  
Substrate Concentrations

The effects of the reaction medium and substrate concentration were studied on the selectivity of Novozym 435 using the asymmetric hydrolysis of dimethyl-3-phenylglutarate as a model reaction. Results show that the use of choline chloride ChCl:urea/phosphate buffer 50% (v/v) as a reaction medium increased the selectivity of Novozym 435 by 16% (e.e = 88%) with respect to the one in 100% phosphate buffer (e.e = 76%). Best results were obtained when high substrate concentrations (well above the solubility limit, 27-fold) and ChCl:urea/phosphate buffer 50% (v/v) as reaction medium at pH 7 and 30 °C were used. Under such conditions, the R-monoester was produced with an enantiomeric purity of 99%. Novozym 435 was more stable in ChCl:urea/phosphate buffer 50% (v/v) than in phosphate buffer, retaining a 50% of its initial activity after 27 h of incubation at pH 7 and 40 °C. Results suggest that the use of deep eutectic solvents (ChCl:urea/phosphate buffer) in an heterogeneous reaction system (high substrate concentration) is a viable and promising strategy for the synthesis of chiral drugs from highly hydrophobic substrates.

scholarly journals Enzymatic hydrolysis of extruded soybean meal at high substrate concentrations

2016 ◽  
pp. 63-73
Author(s):  
Anton Sharikov ◽  
Anna Sereda ◽  
Elena Kostyleva ◽  
Irina Velikoretskaya ◽  
Victor Polyakov
Keyword(s):  
Enzymatic Hydrolysis ◽  
Soybean Meal ◽  
Substrate Concentration ◽  
Degree Of Hydrolysis ◽  
High Substrate ◽  
Bacterial Protease ◽  
Biotechnological Processes ◽  
Enzyme Dosage ◽  
Hydrolysis Of ◽  
Substrate Concentrations

Extrusion as a pretreatment before enzymatic hydrolysis of soybean meal is an effective technique to eliminate antinutritional properties of the main thermostable soy proteins glycinin and ?-conglycinin for production of feed ingredients with enhanced properties. In terms of economic efficiency, biotechnological processes are preferable to carry out at high substrate concentrations. The aim of the investigation was to evaluate the influence of high substrate concentrations in the range of 26-32% and enzyme dosages (0.4 - 3.1 PU/g) on efficiency of hydrolysis of extruded toasted soybean meal with bacterial protease. The results showed that maximum degree of hydrolysis was 42.1% at the enzyme dosage of 3.6 PU/g and at the substrate concentration of 29.0%. The increase in the substrate concentration had a strong effect on the deterioration of dynamic viscosity of the hydrolysates from 0.2 to 5.82 Pa?s. A combination of extrusion cooking at 120?C and enzymatic treatment with ?Protolad B? protease enabled hydrolysis of glycinin and ?-conglycinin to peptides with molecular mass below 15 kDa.

Study on the Process of the Reaction Catalyzed by Polyphenol Oxidase from Purple Sweet Potato

2014 ◽  
Vol 898 ◽  
pp. 153-156 ◽  
Author(s):  
Lu Gao ◽  
Li Chun Zhao ◽  
Ji Dong Duan ◽  
Yan Li Tao
Keyword(s):  
Sweet Potato ◽  
Polyphenol Oxidase ◽  
Substrate Concentration ◽  
Phosphate Buffer ◽  
Phosphate Buffer Solution ◽  
Enzyme Concentration ◽  
Buffer Solution ◽  
Purple Sweet Potato ◽  
The Relationship ◽  
Substrate Concentrations

The polyphenol oxidase (PPO) was extracted from fresh purple sweet potato (PSP) by phosphate buffer solution, and spectrophotometry method was applied in the experiment. The process of the reaction catalyzed by PPO with different substrate concentrations and the relationship between enzyme concentrations and PPO activity were mainly studied here. The result showed that the effect of enzyme concentration on PPO activity was stronger than that of substrate concentration on PPO activity.

scholarly journals Substrate Inhibition in the Hydrolysis of Hippuric Acid Esters by Carboxypeptidase A

1975 ◽  
Vol 53 (2) ◽  
pp. 283-294 ◽  
Author(s):  
Joe Murphy ◽  
John W. Bunting
Keyword(s):  
Substrate Concentration ◽  
Hippuric Acid ◽  
Substrate Inhibition ◽  
Competitive Inhibitor ◽  
Side Chain ◽  
Carboxypeptidase A ◽  
Substrate Activation ◽  
Hydrolysis Of ◽  
Substrate Concentrations ◽  
Acid Esters

The dependence of initial velocity upon substrate concentration has been examined in the carboxypeptidase A catalyzed hydrolysis of the following hippuric acid esters (at pH 7.5, 25°, ionic strength O.5): C6H5CONHCH2CO2CHRCO2H: R=CH3; CH2CH3;(CH2)2CH3; (CH2)3CH3; (CH2)5CH3; CH(CH3)2; CH2CH(CH3)2; C6H5; CH2C6H5. All of these esters display marked substrate inhibition of their enzymic hydrolyses. With the exception of R=CH3, the velocity-substrate concentration profiles for each of these esters can be rationalized by the formation of an E.S2 complex which, independent of the alcohol moiety of the ester, reacts approximately 25 times more slowly than the E.S complex. For most of these esters, the formation of E.S2 approximates ordered binding of the substrate molecules at the catalytic and inhibitory sites. While binding at the catalytic site is markedly dependent on the nature of the R group, binding of a second substrate molecule to E.S is not significantly affected by the nature of the R side chain. For R=C6H5, the D ester is neither a substrate nor a competitive inhibitor of the hydrolysis of the L-ester but can replace the L-ester at the binding site which is responsible for substrate inhibition. The kinetic analysis suggests that this behavior of D and L -enantiomers is also typical of the other esters examined (except possibly R=CH3). For R=CH3 only, substrate activation also seems to occur prior to the onset of substrate inhibition at higher substrate concentrations.

scholarly journals Synthesis of 5-Hydroxymethylfurfural from Cassava (Manihot utilissima pohl) Peels through Dehydration Reaction using Deep Eutectic Solvent Based on Choline Chloride/Citric Acid

2021 ◽  
Vol 33 (5) ◽  
pp. 1115-1119
Author(s):  
R. Manurung ◽  
H. Silalahi ◽  
O. Winda ◽  
A.G. Siregar
Keyword(s):  
Citric Acid ◽  
Reaction Temperature ◽  
Choline Chloride ◽  
Deep Eutectic Solvent ◽  
Deep Eutectic Solvents ◽  
Cellulose Content ◽  
Dehydration Reaction ◽  
Reaction Conditions ◽  
Hydrolysis Of ◽  
Cassava Peel

The high cellulose content in cassava peel has an opportunity to produce bio-based chemical products in 5-hydroxymethylfurfural (5-HMF) form. This study aimed to determine the optimum conditions of glucose dehydration reaction as a result of hydrolysis of the best cassava peel cellulose. The variables observed in this study were H2SO4 catalyst concentrations in the hydrolysis reaction, temperature and amount of deep eutectic solvents based on choline chloride/citric acid. The optimum dehydration reaction conditions in this study was the glucose:deep eutectic solvents mass ratio of 1:6 at the reaction temperature of 80 ºC. The highest yield of 64.50% at an initial glucose concentration of 5.70% using a 1.5% H2SO4 catalyst during the hydrolysis of cassava peel cellulose. The results obtained in this study indicated that addition of choline chloride/citric acid as deep eutectic solvent can increase the yield of 5-HMF.

scholarly journals Using Choline Chloride-Based DESs as Co-Solvent for 3,5-Bis(trifluoromethyl) Acetophenone Bioreduction with Rhodococcus erythropolis XS1012

Catalysts ◽  
2019 ◽  
Vol 10 (1) ◽  
pp. 30 ◽  
Author(s):  
Chen ◽  
Qian ◽  
Lin ◽  
Chen ◽  
Wang
Keyword(s):  
Substrate Concentration ◽  
Choline Chloride ◽  
Cell Activity ◽  
Great Influence ◽  
Rhodococcus Erythropolis ◽  
Deep Eutectic Solvents ◽  
Cell Membrane Permeability ◽  
Buffer System ◽  
High Substrate Concentration ◽  
Reaction Conditions

(S)-3,5-Bistrifluoromethylphenyl ethanol((S)-BTPE) is a key pharmaceutical intermediate of the NK-1 receptor antagonist. The asymmetric bioreduction of 3,5-bis(trifluoromethyl) acetophenone (BTAP) to (S)-BTPE using Rhodococcus erythropolis XS1012 has been established in a phosphate buffer system. To overcome the problem of unsatisfactory yields at high substrate concentration, deep eutectic solvents (DESs) have been introduced to the buffer system. After screening 13 kinds of choline chloride-based DESs, [choline chloride][urea] ([ChCl][U]) showed great influence on the cell activity and significantly increased the cell membrane permeability. Subsequently, some major parameters for this reaction were determined. A remarkable (S)-BTPE yield of 91.9% was gained at 150 mM substrate concentration under optimized reaction conditions with >99.9% product enantioselectivity. Compared to reduction in a buffer system, the developed [ChCl][U]-containing system increased the yield from 82.6% to 91.9%. It maintains a yield of 80.7% with the substrate concentration up to 300 mM, compared to only 63.0% in buffer system. This study demonstrated that [ChCl][U] is a feasible co-solvent to improve the bioreduction process.

Kinetic laws for solid-supported enzymes

1970 ◽  
Vol 48 (10) ◽  
pp. 1498-1504 ◽  
Author(s):  
P. V. Sundaram ◽  
A. Tweedale ◽  
K. J. Laidler
Keyword(s):  
Substrate Concentration ◽  
Free Solution ◽  
Kinetic Behavior ◽  
Michaelis Constant ◽  
Solid Supports ◽  
High Substrate ◽  
Substrate Concentrations

Enzymes behave differently when attached to solid supports for four main reasons: (1) their conformations when they are supported may differ from those in free solution, (2) they act upon substrates in a different environment, (3) there will be partitioning of substrate between the support and the free solution, and (4) there will be effects due to diffusion of the substrate in the support. The present paper examines effects (3) and (4) and shows how rates will vary with substrate concentration. If factors (1) and (2) do not enter, rates in the limit of high substrate concentrations will be the same for the supported enzyme as in free solution. At low substrate concentrations, rates will be less for the supported enzyme if the substrate is less soluble in the support than in free solution, and the apparent Michaelis constant, Km(app.), will be greater; conversely, for higher solubility in the support, rates will be greater and Km(app.) smaller. Effect (4) leads to lower rates and higher Km(app.) values, except in the limit of high substrate concentrations. At a sufficiently low thickness of the support, depending upon the activity of the enzyme, the kinetic behavior becomes identical with that in free solution.

scholarly journals DOES THE KINETICS OF TRYPSIN DIGESTION DEPEND ON THE FORMATION OF A COMPOUND BETWEEN ENZYME AND SUBSTRATE

1922 ◽  
Vol 4 (5) ◽  
pp. 487-509 ◽  
Author(s):  
John H. Northrop
Keyword(s):  
Experimental Evidence ◽  
Substrate Concentration ◽  
Relative Velocity ◽  
Mass Action ◽  
Law Of Mass Action ◽  
Rate Of Hydrolysis ◽  
Gelatin Concentration ◽  
Kinetics Of ◽  
Hydrolysis Of ◽  
Substrate Concentrations

1. The velocity of hydrolysis of gelatin by trypsin increases more slowly than the gelatin concentration and finally becomes nearly independent of the gelatin concentration. The relative velocity of hydrolysis of any two substrate concentrations is independent of the quantity of enzyme used to make the comparison. 2. The rate of hydrolysis is independent of the viscosity of the solution. 3. The percentage retardation of the rate of hydrolysis by inhibiting substances, is independent of the substrate concentration. 4. There is experimental evidence that the enzyme and inhibiting substance are combined to form a widely dissociated compound. 5. If the substrate were also combined with the enzyme, an increase in the substrate concentration should affect the equilibrium between the enzyme and the inhibiting substance. This is not the case. 6. The rate of digestion of a mixture of casein and gelatin is equal to the sum of the rates of hydrolysis of the two substances alone, as it should be if the rate is proportional to the concentration of free enzyme. This contradicts the saturation hypothesis. 7. If the reaction is followed by determining directly the change in the substrate concentration, it is found that this change agrees with the law of mass action; i.e., the rate of digestion is proportional to the substrate concentration.

Flow Kinetics of β-Galactosidase Chemically Attached to Nylon Tubing

1975 ◽  
Vol 53 (10) ◽  
pp. 1061-1069 ◽  
Author(s):  
D. Narinesingh ◽  
T. T. Ngo ◽  
K. J. Laidler
Keyword(s):  
Flow Rate ◽  
Substrate Concentration ◽  
Diffusion Control ◽  
Theoretical Treatment ◽  
Enzyme Reactors ◽  
Diffusion Controlled ◽  
Nylon Tubing ◽  
Kinetics Of ◽  
Hydrolysis Of ◽  
Substrate Concentrations

β-Galactosidase (EC 3.2.1.23) has been attached covalently to the inner surface of nylon tubing. An experimental study has been made of the flow kinetics for the hydrolysis of o-nitrophenylgalactose, the substrate concentration and flow rate being varied. The results were analyzed in the light of the theoretical treatment of Kobayashi and Laidler, three different methods of analysis being employed. It is concluded that at the lower substrate concentrations and flow rates employed, the reactions are largely diffusion controlled; with increase in flow rate and substrate concentration the width of the Nernst diffusion layer decreases, and there is found to be less diffusion control. The values of Km(app) vary with flow rate VF, being linear in VF−1/3, and the value extrapolated to very high flow rate agrees well with the Km value for β-galactosidase in free solution. The theory and results are shown to provide guidelines for the design of open tubular heterogeneous enzyme reactors for industrial, biomedical, and analytical applications.

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