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Related Concept Videos

Calculating pH Changes in a Buffer Solution02:45

Calculating pH Changes in a Buffer Solution

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A buffer can prevent a sudden drop or increase in the pH of a solution after the addition of a strong acid or base up to its buffering capacity; however, such addition of a strong acid or base does result in the slight pH change of the solution. The small pH change can be calculated by determining the resulting change in the concentration of buffer components, i.e., a weak acid and its conjugate base or vice versa. The concentrations obtained using these stoichiometric calculations can be used...
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Buffers02:56

Buffers

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A solution containing appreciable amounts of a weak conjugate acid-base pair is called a buffer solution, or a buffer. Buffer solutions resist a change in pH when small amounts of a strong acid or a strong base are added. A solution of acetic acid and sodium acetate is an example of a buffer that consists of a weak acid and its salt: CH3COOH (aq) + CH3COONa (aq). An example of a buffer that consists of a weak base and its salt is a solution of ammonia and ammonium chloride: NH3 (aq) + NH4Cl...
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Buffers: Buffer Capacity01:09

Buffers: Buffer Capacity

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Buffer capacity is the quantitative measure of a buffer to resist the change in pH. As shown in the following equation, the buffer capacity, denoted by 'beta', is expressed as the number of moles of acid or base needed to change the pH of a one-liter buffer solution by 1 unit. Here, Ca and Cb indicate the number of moles of acid and base, respectively. Note that dpH represents the change in pH.
In the graph, pH is plotted as a function of the number of moles of base (Cb) added to a weak...
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Buffer Effectiveness02:19

Buffer Effectiveness

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Buffer solutions do not have an unlimited capacity to keep the pH relatively constant . Instead, the ability of a buffer solution to resist changes in pH relies on the presence of appreciable amounts of its conjugate weak acid-base pair. When enough strong acid or base is added to substantially lower the concentration of either member of the buffer pair, the buffering action within the solution is compromised.
The buffer capacity is the amount of acid or base that can be added to a given volume...
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Enzymes02:34

Enzymes

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Inside living organisms, enzymes act as catalysts for many biochemical reactions involved in cellular metabolism. The role of enzymes is to reduce the activation energies of biochemical reactions by forming complexes with its substrates. The lowering of activation energies favor an increase in the rates of biochemical reactions.
Enzyme deficiencies can often translate into life-threatening diseases. For example, a genetic abnormality resulting in the deficiency of the enzyme G6PD...
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Phosphate Buffer01:22

Phosphate Buffer

5.2K
The phosphate buffer system is a critical biological mechanism for maintaining pH stability in the body. This system operates primarily through two components: sodium dihydrogen phosphate (NaH2PO4), which acts as a weak acid, and sodium hydrogen phosphate (Na2HPO4), which serves as a weak base.
Sodium dihydrogen phosphate does not fully dissociate in neutral or acidic solutions. When a strong base, such as sodium hydroxide (NaOH), is introduced into the solution, sodium dihydrogen phosphate...
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Related Experiment Video

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High-throughput Fluorometric Measurement of Potential Soil Extracellular Enzyme Activities
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A non-specific nucleolytic enzyme and its application potential in EDTA-containing buffer solutions.

Sarah Schmitz1, Volker Nölle1, Skander Elleuche2

  • 1Miltenyi Biotec GmbH, Friedrich-Ebert-Straße 68, 51429, Bergisch Gladbach, Germany.

Biotechnology Letters
|November 4, 2018
PubMed
Summary

A novel enzyme, EcNuc, from Escherichia coli functions without metal ions. This non-specific nuclease effectively degrades nucleic acids in EDTA-containing buffers, aiding protein purification.

Keywords:
Biotechnological applicationDNaseEDTARNase

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Area of Science:

  • Biochemistry
  • Molecular Biology

Background:

  • Metal-ion independent non-specific nucleases are valuable for bioprocessing.
  • EDTA is commonly used in bioprocessing, often chelating essential metal ions.

Purpose of the Study:

  • To identify and characterize a novel metal-ion independent non-specific nuclease.
  • To evaluate its potential application in EDTA-containing bioprocessing workflows.

Main Methods:

  • Identification and cloning of the EcNuc gene from Escherichia coli.
  • Expression and purification of recombinant EcNuc protein.
  • Enzyme activity assays under varying conditions (temperature, pH, EDTA, metal ions).

Main Results:

  • A novel extracellular non-specific nuclease, EcNuc, was identified and characterized.
  • EcNuc exhibits optimal activity at 41.7°C and pH 5.8.
  • EcNuc tolerates up to 20 mM EDTA and is not inhibited by high metal ion concentrations in the absence of EDTA.
  • EcNuc reduces the viscosity of crude protein extracts lysed in EDTA-containing buffers.

Conclusions:

  • EcNuc is a metal-ion independent non-specific nuclease with potential for bioprocessing.
  • Proof-of-concept demonstrated its utility for nucleic acid removal in EDTA-containing buffers for protein purification.