Use of chelating agents

Chelating agents are molecules that have the ability to form more than one bond to a metal ion, thereby increasing the stability of the ion complex.

The complex is called a chelate, which is a word from the Greek chela, meaning “claw”

Naturally occurring chelates These are necessary for many life-

sustaining processes. The vitamin B12-cobalt chelate complex is a dietary requirement in all higher animals.

Chlorophyll-magnesium complex is a critical component of photosynthesis.

The hemoglobin-iron complex transports oxygen through the blood.

Chelates in industry

Chelating agents have found uses in industry as detergents, additives, stabilizing agents, preservatives, and flavor and color retainers. Ethylenediaminetetraacetic acid (EDTA) is a quadridentate or hexadentate chelating agent; it is capable of forming either four or six bonds with metal ions.

EDTA

EDTA is widely used for enhancing the cleaning power of detergents and soaps by forming chelates with the magnesium and calcium metals in hard water.

Chelating agents are also used as color retainers for textile dyes.

EDTA “ cont”

EDTA is also used as a food additive. Food spoilage is catalyzed by enzymes that require certain metals in order to work. Because of its ability to sequester these metal ions, EDTA deactivates these enzymes that would otherwise cause the food products to become rancid.

EDTA also deactivates the enzymes that cause food to lose color or flavor.

EDTA “ cont”

In medicine, EDTA is used to preserve blood by sequestering the calcium ions that cause clotting. Administered intravenously, EDTA is effective in treating cadmium, lead, and iron poisoning by chelating the metal ions and removing them from the blood stream.

The sequestered metals are removed from the blood stream and typically eliminated in the urine.

Role of EDTA in stabilization of samples EDTA at a final concentration

of 1-5 mM avoid metal-induced oxidation of –SH groups of protein samples and helps to maintain the protein in a reduced state.

Purified proteins often need to be stored for an extended period of time while retaining their original structural integrity and/or activity.

The extent of storage ‘shelf life’ can vary from a few days to more than a year and is dependent on the nature of the protein and the storage conditions used.

Optimal conditions for storage are distinctive to each protein; nevertheless, it is possible to suggest some general guidelines for protein storage and stability.

General Considerations for Protein Storage Generally, proteins are best stored at

≤ 4°C in clean, autoclaved glassware or polypropylene tubes.

Storage at room temperature often leads to protein degradation and/or inactivity, commonly as a result of microbial growth.

Additives

Many compounds may be added to protein solutions to lengthen shelf life:

Cryoprotectants such as glycerol or ethylene glycol to a final concentration of 25-50% help to stabilize proteins by preventing the formation of ice crystals at -20°C that destroy protein structure.

Protease inhibitors prevent proteolytic cleavage of proteins .

Anti-microbial agents such as sodium azide (NaN3) at a final concentration of 0.02-0.05% (w/v) or thimerosal at a final concentration of 0.01 % (w/v) inhibit microbial growth

Reducing agents such a dithiothreitol (DTT) and 2-mercaptoethanol (2-ME) at final concentrations of 1-5 mM also help to maintain the protein in the reduced state by preventing oxidation of cysteines.

DNA extraction often results in the recovery of low concentration solutions of DNA that are prone to DNA degradation or other loss.

It is critical that optimal methods are employed for their long-term storage.

Storage temepratures

DNA can be stored in different temperatures :

(+4°C, -20°C, -80°C) However, the highest quantity of

DNA remained in samples stored at -80°C.

Storage additives

Three additives (Tris , EDTA (TE) buffer, trehalose).

The highest quantity of DNA remained in samples stored at -80 degrees C, regardless of storage additives, DNA quality was best preserved in the presence of trehalose which is an alpha-linked disaccharide formed by an α,α-1,1-glucoside bond between two α-glucose units.