Nonactin and other nactines

This webpage has been created by the students of the OpenUniversity.

This page contains information taken from Wikipedia, PUBMED, wendors pages, and our own laboratory work under the supervision of Dr AbuAmir Mandallah.

About Nonactin

[from Wikipedia, re-edited]

Nonactin is a member of a family of naturally occurring cyclic ionophores known as the macrotetrolide antibiotics. The other members of this homologous family are monactin, dinactin, trinactin and tetranactin which are all neutral ionophoric substances and higher homologs of nonactin. Collectively, this class is known as the Nactins.

It was isolated by Corbaz et al. in 1955 from bacterial strains.

Nonactin is known for its ability to form complexes with alkali cations, most notably potassium and sodium. It also displays a marked binding preference for potassium ion over sodium ion. In general, nonactin (and other members of the nactin family) exhibits binding preferences for some ions over others. This ion selectivity is seen in other macrocyclic ligands such as the cyclic ionophore valinomycin which is also an antibiotic. Ion selectivity is also observed in crown ethers which also exhibit a discriminatory preference of some ions over others. Although nonactin (and in fact, all nactins) exhibits an especially high cation selectivity for potassium ion over sodium and rubidium ion, it exhibits the highest selectivity for ammonium ion and thallium ion.

During complexation, the nonactin backbone convolutes into a pattern resembling the seam of a tennis ball. In the K+-nonactin complex, the the K+ ion is entirely surrounded by four carbonyl oxygen atoms and the four oxygen atoms of the tetrahydrofuran ring. These eight oxygen atoms that surround the ion are nearly equidistant from it and adopt a nearly cubic coordination sphere around the ion. In this complex, all polar carbonyl groups point inwards and nonpolar moieties point outwards, thus building up a hydrophobic exterior for the complex and making it soluble in lipid membranes. This is how nonactin is able to transport K+ ions across lipid membranes.

Nonactin has been reported to specifically inhibit the processing of cytoplasmic precursor proteins destined for the mitochondria.It is able to uncouple the oxidative phosphorylation of mitochondria of rat liver in a low concentration, and can also carry cations across biological and artificial membranes.

Nactines are also used as ammonium ion sensors in membrane electrodes and as pesticides. Nonactin is soluble in Methanol, Dichloromethane, Ethyl Acetate and DMSO (Dimethyl sulfoxide). It is insoluble in water.

[from Dr Abu-Amir Mandallah course]

It is unclear whether the biological activity of nonactin (and other nactines) is related to its ionophoric property. It is says in the literature that the biologic activity of nactines grows as their "number" grows, and nonactin is the least potent, nearly inactive, and the tetranactin is the strongest one. However it is clear that all 5 compounds have activity similar in properties, and differ in potency.

There are publications about making nonactin for agricultural use. Because Nonactin is said to be the least potent of nactines, an effort was made to get best yield of tetranactin in the fermentation. The best result was obtained using streptomyces griseus, and adding a lot of Phosphate and organic acids such as acetic, to the fermentation medium. Actually, the commercial preparation called "Polynactin" (not available any more) purposely contained no nonactin at all.

Some history:

Sigma Aldrich used to make nonactin from streptomyces tsusimaensis, as a byproduct at the Valinomycin fermentation. To test it, they used TLC, normal phase, 6:3:1 Toluene: Ethyl-Acetate: Formic acid. We have found that this TLC puts all nactines in the same spot, and cannot distinguish between the homologs. The "nonactin" from Sigma actually contained no more than 10% nonactin within total of some 98% tetromacrolides. Later Sigma stopped in-house production of Nonactin, and bought it elsewhere. Currently, it seems that Sigma do not make their own analysis on this nonactin, and rely fully on the supplier's QC.

Who makes nonactin

Currently we could identify only three primary makers on Nonactin: Promiliad, Fermentek and American Biodiagnostics.

Both Promiliad and American Biodiagnostics boast to manufacture highly purified nonactin. Promiliad offers two products, pure and extrapure, the last being three times more expensive. Fermentek is working on their own version of "extra pure Nonactin", and promise it will be world's both best and cheapest.

Currently (2006), it seems that the only usage for nonactin is the specific electrodes manufacturing. Professor Priestley, renown as Nonactin guru, explains that the Nonactin used for ammonium electrodes, should better be pure, lest the calibration of these electrodes would be more difficult, but it has not be proven that other preparations, such as tetranactin, or mixture of nactines might not be used.

Our estimations are that if a tetromacrolide mixture could be used it might be as cheap as $20 / 1 gram.

About the analysis of Nonactin and Nactines.

[ from promiliad website ]

"As TLC is a notoriously unreliable method for estimating the nonactin content of macrotetrolide mixtures, we measure nonactin content by proton nuclear magnetic resonance spectroscopy. This technique allows us to accurately quantify the proportions of individual homologues in our products."

[ from American bio diagnostics website]

The measurement of the nonactin content requires care. TLC measurements are wholly unreliable. Each of the components stains to a different extent; size.

[our own observations]

Not only that the spots of the various nactines vary in size and coloration intensity, but even their very RF is not stable. The greater the concentration, the higher the spot appears on the TLC plate.

In the scientific publications we found a lot of usage of HPLC analysis, employing nonstandard detectors, such as light scattering, etc. Nobody had used simple UV detector. Therefore we had trouble to reproduce these measurements. Nor could we establish the spectra of the higher homologues.

It seems that currently, all manufacturers are using NMR analysis for this product.