In Silico Investigation of Luminol, Its Analogues and Mechanism of Chemiluminescence for Blood Identification Beyond Forensics

Abstract

This study aimed at discovering chemiluminescent analogues of luminol, predict their molecular binding to hemoglobin of bloodstains in household crime, and expound the mechanism of chemiluminescence of luminol. Similarity and clustering analyses of luminol analogues were conducted, and molecular docking was carried out using hemoglobin from Homo sapiens and four domestic organisms namely Gallus gallus, Drosophila melanogaster, Rattus norvegicus, and Canis familiaris. The results showed the order of overall binding score as D. melanogaster > H. sapiens > C. familiaris > R. norvegicus > G. gallus. Seven compounds namely ZINC16958228, ZINC17023010, ZINC19915427, ZINC34928954, ZINC19915369, ZINC19915444, and ZINC82294978, were found to be consistently stable in binding with diverse hemoglobin and possibly have chemiluminescence than luminol in this in silico study. The interaction of human hemoglobin with luminol and its analogues, showed that amino acid residues His45, Lys61, Asn68, Val73, Met76, Pro77, Ala79, Ala82, Leu83, Pro95, Phe98, Lys99, Ser102, Ser133, Ala134, and Thr134, were possibly significant in the mechanism of action of presumptive test compounds. It was hypothesized that the improved mechanism of chemiluminescent for the identification of blood was based on peroxidase-like reaction, that produces nitric oxide which binds to hemoglobin (Hb) and inhibits Hb degradation without yielding fluorescent products. The compound 2,3-benzodioxine-1,4,5(6H)-trione was formed, which possibly emits light. This study provides novel insight on the luminol and its expanded mechanism for broader possible applications with careful development of new methodologies.