Nucleophilic addition to the 9 position of 9‐phenylcarboxylate‐10‐methylacridinium protects against hydrolysis of the ester

Abstract

The chemiluminescent reaction of an acridinium ester (AE) requires addition of peroxide to the 9 position of the acridinium ring. The addition of a hydroxide ion to the 9 position of an acridinium ester to form the carbinol adduct has also been well documented. We have observed a similar addition of other nucleophiles to the acridinium ring to form an acridan adduct. The adduct formed with bisulphite has been particularly well-characterized for rate of formation, rate of reversion, and reaction equilibrium. The formation of an adduct (other than H2O2) has been demonstrated to decrease significantly the reactivity of the adjacent ester bond to alkaline hydrolysis. The resulting, more stable adduct is very useful when the acridinium ester is used as a label in DNA probe-based assays. The adduct is highly resistant to hydrolysis under the conditions often desired for DNA probe-based assays (high temperature, elevated pH, extended storage).

Employment of a Phenoxy-Substituted Acridinium Ester as a Long-Lived Chemiluminescent Indicator of Glucose Oxidase Activity and Its Application in an Alkaline Phosphatase Amplification Cascade Immunoassay

“…It was found necessary to add the di-ortho-bromo-AE to the reaction buffer within a short time (10 min) prior to the addition of the GOx, otherwise the resultant yield was reduced. This may have been due to nonchemiluminescent adduct and/or pseudobase formation when the AE was removed from its acidic storage environment (7). Such phenomena may have been occurring during the incubation and may have been partially responsible for the fall in output seen at high GOx concentration in addition to that resulting from peroxide-dependent consumption of AE.…”

Kinetics and mechanism of covalent addition of S(IV) species to the acridinium cation

“…Of the three acridine species, only the acridinium cation 1 reacts and the quaternary ring nitrogen was found to be essential for adduct formation (12), with the resulting loss of ring aromaticity. We did not obtain any kinetic information in basic solutions (pH > 8) due to pseudobase formation and the small value of kob,,.…”

“…has been extensively studied. Pitmann and Sternson (1 1) estimated the equilibrium constant of the addition of the sulfite ion to the acridinium cation at 25°C and ionic strength I = 1.0 M and Hammond et al (12) studied the addition of bisulfite to the 9-phenylcarboxylate-10-methylacridinium cation from the equilibrium point of view and describe some aspects of the macroscopic rate of the process. In any case, there is no information about the kinetics, mechanism, or any other factors that might affect the formation of adduct acridine-S(1V).…”

Chemiluminescence Energy Transfer Processes and Micellar Effects

“…The acridinium ester used was 2,4,6-trichlorophenyl-10-methylacridinium-9-carboxylate fluorosulfonate (synthesized by T. J. Novak 28 ) and was stored as a 1 mM solution in acetonitrile. A working solution was made by dilution in water which had been adjusted to pH 3 with nitric acid to prevent hydrolysis . A buffer of 0.1 M phosphate pH 9.0 was prepared using potassium phosphate, dibasic, and dilute phosphoric acid.…”